Publications by authors named "Alan Davidson"

254 Publications

The systemic inflammatory response and clinicopathological characteristics in patients admitted to hospital with COVID-19 infection: Comparison of 2 consecutive cohorts.

PLoS One 2021 27;16(5):e0251924. Epub 2021 May 27.

Academic Unit of Surgery, School of Medicine, University of Glasgow, New Lister Building, Royal Infirmary, Glasgow, United Kingdom.

Background: In order to manage the COVID-19 systemic inflammatory response, it is important to identify clinicopathological characteristics across multiple cohorts.

Methods: The aim of the present study was to compare the 4C mortality score, other measures of the systemic inflammatory response and clinicopathological characteristics in two consecutive cohorts of patients on admission with COVID-19. Electronic patient records for 2 consecutive cohorts of patients admitted to two urban teaching hospitals with COVID-19 during two 7-week periods of the COVID-19 pandemic in Glasgow, U.K. (cohort 1: 17/3/2020-1/5/2020) and (cohort 2: 18/5/2020-6/7/2020) were examined for routine clinical, laboratory and clinical outcome data.

Results: Compared with cohort 1, cohort 2 were older (p<0.001), more likely to be female (p<0.05) and have less independent living circumstances (p<0.001). More patients in cohort 2 were PCR positive, CXR negative (both p<0.001) and had low serum albumin concentrations (p<0.001). 30-day mortality was similar between both cohorts (23% and 22%). In cohort 2, age >70 (p<0.05), male gender (p<0.05), COPD (p<0.05), cognitive impairment (p<0.05), frailty (p<0.001), delirium (p = 0.001), CRP>150mg/L (p<0.05), albumin <30 g/L (p<0.01), elevated perioperative Glasgow Prognostic Score (p<0.05), elevated neutrophil-lymphocyte ratio (p<0.001), low haematocrit (p<0.01), elevated PT (p<0.05), sodium <133 mmol/L (p<0.01) elevated urea (p<0.001), creatinine (p<0.001), glucose (p<0.05) and lactate (p<0.001) and the 4C score (p<0.001) were associated with 30-day mortality. In multivariate analysis, greater frailty (CFS>3) (OR 11.3, 95% C.I. 2.3-96.7, p<0.05), low albumin (<30g/L) (OR 2.5, 95% C.I. 1.0-6.2, p<0.05), high NLR (≥3) (OR 2.2, 95% C.I. 1.5-4.5, p<0.05) and the 4C score (OR 2.4, 95% C.I. 1.0-5.6, p<0.05) remained independently associated with 30-day mortality.

Conclusion: In addition to the 4C mortality score, frailty score and a low albumin were strongly independently associated with 30-day mortality in two consecutive cohorts of patients admitted to hospital with COVID-19.

Trial Registration: clinicaltrials.gov: NCT04484545.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0251924PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8159002PMC
June 2021

A Simplified Method for Generating Kidney Organoids from Human Pluripotent Stem Cells.

J Vis Exp 2021 04 13(170). Epub 2021 Apr 13.

Department of Developmental Biology, University of Pittsburgh, School of Medicine; Center for Critical Care Nephrology, University of Pittsburgh, School of Medicine;

Kidney organoids generated from hPSCs have provided an unlimited source of renal tissue. Human kidney organoids are an invaluable tool for studying kidney disease and injury, developing cell-based therapies, and testing new therapeutics. For such applications, large numbers of uniform organoids and highly reproducible assays are needed. We have built upon our previously published kidney organoid protocol to improve the overall health of the organoids. This simple, robust 3D protocol involves the formation of uniform embryoid bodies in minimum component medium containing lipids, insulin-transferrin-selenium-ethanolamine supplement and polyvinyl alcohol with GSK3 inhibitor (CHIR99021) for 3 days, followed by culture in knock-out serum replacement (KOSR)-containing medium. In addition, agitating assays allows for reduction in clumping of the embryoid bodies and maintaining a uniform size, which is important for reducing variability between organoids. Overall, the protocol provides a fast, efficient, and cost-effective method for generating large quantities of kidney organoids.
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http://dx.doi.org/10.3791/62452DOI Listing
April 2021

Anti-CRISPR AcrIF9 functions by inducing the CRISPR-Cas complex to bind DNA non-specifically.

Nucleic Acids Res 2021 04;49(6):3381-3393

Department of Biochemistry, University of Toronto, 661 University Ave, Toronto, ON, M5G 1M1, Canada.

Phages and other mobile genetic elements express anti-CRISPR proteins (Acrs) to protect their genomes from destruction by CRISPR-Cas systems. Acrs usually block the ability of CRISPR-Cas systems to bind or cleave their nucleic acid substrates. Here, we investigate an unusual Acr, AcrIF9, that induces a gain-of-function to a type I-F CRISPR-Cas (Csy) complex, causing it to bind strongly to DNA that lacks both a PAM sequence and sequence complementarity. We show that specific and non-specific dsDNA compete for the same site on the Csy:AcrIF9 complex with rapid exchange, but specific ssDNA appears to still bind through complementarity to the CRISPR RNA. Induction of non-specific DNA-binding is a shared property of diverse AcrIF9 homologues. Substitution of a conserved positively charged surface on AcrIF9 abrogated non-specific dsDNA-binding of the Csy:AcrIF9 complex, but specific dsDNA binding was maintained. AcrIF9 mutants with impaired non-specific dsDNA binding activity in vitro displayed a reduced ability to inhibit CRISPR-Cas activity in vivo. We conclude that misdirecting the CRISPR-Cas complex to bind non-specific DNA is a key component of the inhibitory mechanism of AcrIF9. This inhibitory mechanism is distinct from a previously characterized anti-CRISPR, AcrIF1, that sterically blocks DNA-binding, even though AcrIF1and AcrIF9 bind to the same site on the Csy complex.
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http://dx.doi.org/10.1093/nar/gkab092DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8034650PMC
April 2021

The Dynamics of Metabolic Characterization in iPSC-Derived Kidney Organoid Differentiation a Comparative Omics Approach.

Front Genet 2021 10;12:632810. Epub 2021 Feb 10.

CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, China.

The use of differentiating human induced pluripotent stem cells (hiPSCs) in mini-tissue organoids provides an invaluable resource for regenerative medicine applications, particularly in the field of disease modeling. However, most studies using a kidney organoid model, focused solely on the transcriptomics and did not explore mechanisms of regulating kidney organoids related to metabolic effects and maturational phenotype. Here, we applied metabolomics coupled with transcriptomics to investigate the metabolic dynamics and function during kidney organoid differentiation. Not only did we validate the dominant metabolic alteration from glycolysis to oxidative phosphorylation in the iPSC differentiation process but we also showed that glycine, serine, and threonine metabolism had a regulatory role during kidney organoid formation and lineage maturation. Notably, serine had a role in regulating S-adenosylmethionine (SAM) to facilitate kidney organoid formation by altering DNA methylation. Our data revealed that analysis of metabolic characterization broadens our ability to understand phenotype regulation. The utilization of this comparative omics approach, in studying kidney organoid formation, can aid in deciphering unique knowledge about the biological and physiological processes involved in organoid-based disease modeling or drug screening.
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http://dx.doi.org/10.3389/fgene.2021.632810DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7902935PMC
February 2021

Sex Cord Stromal Tumors in Children and Adolescents: A First Report by The South African Children's Cancer Study Group (1990-2015).

J Pediatr Hematol Oncol 2021 Jul;43(5):e619-e624

Division of Paediatric Haematology-Oncology, Chris Hani Baragwanath Academic Hospital, University of Witwatersrand, Soweto, Gauteng.

Objectives: Pediatric sex cord stromal tumors (SCSTs) are extremely rare and there are no reported data from Africa. The authors evaluated the outcomes of children and adolescents with biopsy-proven SCSTs in preparation for the introduction of a national protocol.

Materials And Methods: Retrospective data were collated from 9 South African pediatric oncology units from January 1990 to December 2015. Kaplan-Meier analysis was performed to estimate overall survival (OS) and event-free survival.

Results: Twenty-three patients were diagnosed with SCSTs, 3 male and 20 female individuals, during the study period. Histologies included 1 thecoma, 9 Sertoli-Leydig cell tumors, and 13 juvenile granulosa cell tumors. Stage I tumors predominated (n=14; 60.9%), with 2 stage II (8.7%), 5 stage III (21.7%), and 2 stage IV tumors (8.7%). The upfront resection rate was 91.3% with no reported surgical morbidity or mortality and an OS of 82.1%. Chemotherapy approaches were not standardized. Most children (81.8%), except 2, had recognized platinum-based regimens. Chemotherapy-related toxicity was minimal and acceptable. Assessment of glomerular filtration rate and audiology assessments were infrequent and not standardized. Three patients were lost to follow-up.

Conclusions: Although the numbers in this cohort are small, this study represents the first national cohort in Africa. The 5-year OS of 82.1% was encouraging. Standardized management of rare tumors like SCSTs is critical to improve ensure OS and address potential long-term sequelae.
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http://dx.doi.org/10.1097/MPH.0000000000002076DOI Listing
July 2021

Human Urinal Cell Reprogramming: Synthetic 3D Peptide Hydrogels Enhance Induced Pluripotent Stem Cell Population Homogeneity.

ACS Biomater Sci Eng 2020 11 20;6(11):6263-6275. Epub 2020 Oct 20.

CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou 510530, China.

Somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs), which have promising potential applications in regenerative medicine. However, the challenges of successful applications of human iPSCs for medical purposes are the low generation efficiency, heterogeneous colonies, and exposure to the animal-derived product Matrigel. We aimed to investigate whether human urinal cells could be efficiently reprogrammed into iPSCs in three-dimensional Puramatrix (3D-PM) compared to two-dimensional Matrigel (2D-MG) and to understand how this 3D hydrogel environment affects the reprogramming process. Human urinal cells were successfully reprogrammed into iPSCs in the defined synthetic animal-free 3D-PM. Interestingly, although the colony efficiency in 3D-PM was similar to that in 2D-MG (∼0.05%), the reprogrammed colonies in 3D-PM contained an iPSC population with significantly higher homogeneity, as evidenced by the pluripotent-like morphology and expression of markers. This was further confirmed by transcriptome profile analysis in bulk cells and at the single cell level. Moreover, the homogeneity of the iPSC population in 3D-PM colonies was correlated with the downregulation of integrin β1 (ITGB1) and phosphorylated focal adhesion kinase (FAK). Collectively, 3D-PM provides an alternative approach for obtaining iPSCs with enhanced homogeneity. This work also unveiled the regulation of human somatic cell reprogramming via the extracellular microenvironment.
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http://dx.doi.org/10.1021/acsbiomaterials.0c00667DOI Listing
November 2020

A phage-encoded anti-activator inhibits quorum sensing in Pseudomonas aeruginosa.

Mol Cell 2021 02 6;81(3):571-583.e6. Epub 2021 Jan 6.

Department of Biochemistry, University of Toronto, MaRS West Tower, 661 University Avenue, Toronto, ON M5G 1M1, Canada. Electronic address:

The arms race between bacteria and phages has led to the evolution of diverse anti-phage defenses, several of which are controlled by quorum-sensing pathways. In this work, we characterize a quorum-sensing anti-activator protein, Aqs1, found in Pseudomonas phage DMS3. We show that Aqs1 inhibits LasR, the master regulator of quorum sensing, and present the crystal structure of the Aqs1-LasR complex. The 69-residue Aqs1 protein also inhibits PilB, the type IV pilus assembly ATPase protein, which blocks superinfection by phages that require the pilus for infection. This study highlights the remarkable ability of small phage proteins to bind multiple host proteins and disrupt key biological pathways. As quorum sensing influences various anti-phage defenses, Aqs1 provides a mechanism by which infecting phages might simultaneously dampen multiple defenses. Because quorum-sensing systems are broadly distributed across bacteria, this mechanism of phage counter-defense may play an important role in phage-host evolutionary dynamics.
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http://dx.doi.org/10.1016/j.molcel.2020.12.011DOI Listing
February 2021

Protocol for Large-Scale Production of Kidney Organoids from Human Pluripotent Stem Cells.

STAR Protoc 2020 Dec 29;1(3):100150. Epub 2020 Oct 29.

Department of Molecular Medicine & Pathology, University of Auckland, Auckland 1142, New Zealand.

Kidney organoids represent a physiologically advanced model for studying the mechanisms of kidney development and disease. Here, we describe a simple two-step protocol for the differentiation of human pluripotent stem cells into kidney organoids. Our approach involves suspension culture that allows for rapid and cost-effective bulk production of organoids, which is well suited for large-scale assays such as drug screening. The organoids correspond to fetal human kidney tissue and may be of limited use for modeling adult kidney function. For complete details on the use and execution of this protocol, please refer to Przepiorski et al. (2018).
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http://dx.doi.org/10.1016/j.xpro.2020.100150DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7757290PMC
December 2020

Anti-CRISPR AcrIE2 Binds the Type I-E CRISPR-Cas Complex But Does Not Block DNA Binding.

J Mol Biol 2021 02 16;433(3):166759. Epub 2020 Dec 16.

Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada; Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada. Electronic address:

Anti-CRISPRs are protein inhibitors of CRISPR-Cas systems. They are produced by phages and other mobile genetic elements to evade CRISPR-Cas-mediated destruction. Anti-CRISPRs are remarkably diverse in sequence, structure, and functional mechanism; thus, structural and mechanistic investigations of anti-CRISPRs continue to yield exciting new insights. In this study, we used nuclear magnetic resonance (NMR) spectroscopy to determine the solution structure of AcrIE2, an anti-CRISPR that inhibits the type I-E CRISPR-Cas system of Pseudomonas aeruginosa. Guided by the structure, we used site-directed mutagenesis to identify key residues that are required for AcrIE2 function. Using affinity purification experiments, we found that AcrIE2 binds the type I-E CRISPR-Cas complex (Cascade). In vivo transcriptional assays, in which Cascade was targeted to promoter regions, demonstrated that Cascade still binds to DNA in the presence of AcrIE2. This is the first instance of a type I anti-CRISPR that binds to a CRISPR-Cas complex but does not prevent DNA-binding. Another unusual property of AcrIE2 is that the effect of Cascade:AcrIE2 complex binding to promoter regions varied depending on the position of the binding site. Most surprisingly, Cascade:AcrIE2 binding led to transcriptional activation in some cases rather than repression, which did not occur when Cascade alone bound to the same sites. We conclude that AcrIE2 operates through a distinct mechanism compared to other type I anti-CRISPRs. While AcrIE2 does not prevent Cascade from binding DNA, it likely blocks subsequent recruitment of the Cas3 nuclease to Cascade thereby preventing DNA cleavage.
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http://dx.doi.org/10.1016/j.jmb.2020.166759DOI Listing
February 2021

Phage Proteins Required for Tail Fiber Assembly Also Bind Specifically to the Surface of Host Bacterial Strains.

J Bacteriol 2021 01 11;203(3). Epub 2021 Jan 11.

Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada

To initiate their life cycle, phages must specifically bind to the surface of their bacterial hosts. Long-tailed phages often interact with the cell surface using fibers, which are elongated intertwined trimeric structures. The folding and assembly of these complex structures generally requires the activity of an intra- or intermolecular chaperone protein. Tail fiber assembly (Tfa) proteins are a very large family of proteins that serve as chaperones for fiber folding in a wide variety of phages that infect diverse species. A recent structural study showed that the Tfa protein from phage Mu (Tfa) mediates fiber folding and stays bound to the distal tip of the fiber, becoming a component of the mature phage particle. This finding revealed the potential for Tfa to also play a role in cell surface binding. To address this issue, we have here shown that Tfa binds to lipopolysaccharide (LPS), the cell surface receptor of phage Mu, with a similar strength as to the fiber itself. Furthermore, we have found that Tfa and the Tfa protein from phage P2 bind LPS with distinct specificities that mirror the host specificity of these two phages. By comparing the sequences of these two proteins, which are 93% identical, we identified a single residue that is responsible for their distinct LPS-binding behaviors. Although we have not yet found conditions under which Tfa proteins influence host range, the potential for such a role is now evident, as we have demonstrated their ability to bind LPS in a strain-specific manner. With the growing interest in using phages to combat antibiotic-resistant infections or manipulate the human microbiome, establishing approaches for the modification of phage host range has become an important research topic. Tfa proteins are a large family of proteins known previously to function as chaperones for the folding of phage fibers, which are crucial determinants of host range for long-tailed phages. Here, we reveal that some Tfa proteins are bi-functional, with the additional activity of binding to LPS, the surface binding receptor for many phages. This discovery opens up new potential avenues for altering phage host range through engineering of the surface binding specificity of Tfa proteins.
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http://dx.doi.org/10.1128/JB.00406-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7811200PMC
January 2021

Travel burden and clinical presentation of retinoblastoma: analysis of 1024 patients from 43 African countries and 518 patients from 40 European countries.

Br J Ophthalmol 2020 Sep 15. Epub 2020 Sep 15.

Pediatric Oncology Unit, Hospital Universitario y Politécnico La Fe, Valencia, Spain.

Background: The travel distance from home to a treatment centre, which may impact the stage at diagnosis, has not been investigated for retinoblastoma, the most common childhood eye cancer. We aimed to investigate the travel burden and its impact on clinical presentation in a large sample of patients with retinoblastoma from Africa and Europe.

Methods: A cross-sectional analysis including 518 treatment-naïve patients with retinoblastoma residing in 40 European countries and 1024 treatment-naïve patients with retinoblastoma residing in 43 African countries.

Results: Capture rate was 42.2% of expected patients from Africa and 108.8% from Europe. African patients were older (95% CI -12.4 to -5.4, p<0.001), had fewer cases of familial retinoblastoma (95% CI 2.0 to 5.3, p<0.001) and presented with more advanced disease (95% CI 6.0 to 9.8, p<0.001); 43.4% and 15.4% of Africans had extraocular retinoblastoma and distant metastasis at the time of diagnosis, respectively, compared to 2.9% and 1.0% of the Europeans. To reach a retinoblastoma centre, European patients travelled 421.8 km compared to Africans who travelled 185.7 km (p<0.001). On regression analysis, lower-national income level, African residence and older age (p<0.001), but not travel distance (p=0.19), were risk factors for advanced disease.

Conclusions: Fewer than half the expected number of patients with retinoblastoma presented to African referral centres in 2017, suggesting poor awareness or other barriers to access. Despite the relatively shorter distance travelled by African patients, they presented with later-stage disease. Health education about retinoblastoma is needed for carers and health workers in Africa in order to increase capture rate and promote early referral.
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http://dx.doi.org/10.1136/bjophthalmol-2020-316613DOI Listing
September 2020

Prognostic factors in patients admitted to an urban teaching hospital with COVID-19 infection.

J Transl Med 2020 09 15;18(1):354. Epub 2020 Sep 15.

Academic Unit of Surgery, School of Medicine, University of Glasgow, New Lister Building, Royal Infirmary, Glasgow, G31 2ER, UK.

Background: Severe COVID-19 infection results in a systemic inflammatory response (SIRS). This SIRS response shares similarities to the changes observed during the peri-operative period that are recognised to be associated with the development of multiple organ failure.

Methods: Electronic patient records for patients who were admitted to an urban teaching hospital during the initial 7-week period of the COVID-19 pandemic in Glasgow, U.K. (17th March 2020-1st May 2020) were examined for routine clinical, laboratory and clinical outcome data. Age, sex, BMI and documented evidence of COVID-19 infection at time of discharge or death certification were considered minimal criteria for inclusion.

Results: Of the 224 patients who fulfilled the criteria for inclusion, 52 (23%) had died at 30-days following admission. COVID-19 related respiratory failure (75%) and multiorgan failure (12%) were the commonest causes of death recorded. Age ≥ 70 years (p < 0.001), past medical history of cognitive impairment (p ≤ 0.001), previous delirium (p < 0.001), clinical frailty score > 3 (p < 0.001), hypertension (p < 0.05), heart failure (p < 0.01), national early warning score (NEWS) > 4 (p < 0.01), positive CXR (p < 0.01), and subsequent positive COVID-19 swab (p ≤ 0.001) were associated with 30-day mortality. CRP > 80 mg/L (p < 0.05), albumin < 35 g/L (p < 0.05), peri-operative Glasgow Prognostic Score (poGPS) (p < 0.05), lymphocytes < 1.5 10/l (p < 0.05), neutrophil lymphocyte ratio (p ≤ 0.001), haematocrit (< 0.40 L/L (male)/ < 0.37 L/L (female)) (p ≤ 0.01), urea > 7.5 mmol/L (p < 0.001), creatinine > 130 mmol/L (p < 0.05) and elevated urea: albumin ratio (< 0.001) were also associated with 30-day mortality. On multivariate analysis, age ≥ 70 years (O.R. 3.9, 95% C.I. 1.4-8.2, p < 0.001), past medical history of heart failure (O.R. 3.3, 95% C.I. 1.2-19.3, p < 0.05), NEWS > 4 (O.R. 2.4, 95% C.I. 1.1-4.4, p < 0.05), positive initial CXR (O.R. 0.4, 95% C.I. 0.2-0.9, p < 0.05) and poGPS (O.R. 2.3, 95% C.I. 1.1-4.4, p < 0.05) remained independently associated with 30-day mortality. Among those patients who tested PCR COVID-19 positive (n = 122), age ≥ 70 years (O.R. 4.7, 95% C.I. 2.0-11.3, p < 0.001), past medical history of heart failure (O.R. 4.4, 95% C.I. 1.2-20.5, p < 0.05) and poGPS (O.R. 2.4, 95% C.I. 1.1-5.1, p < 0.05) remained independently associated with 30-days mortality.

Conclusion: Age ≥ 70 years and severe systemic inflammation as measured by the peri-operative Glasgow Prognostic Score are independently associated with 30-day mortality among patients admitted to hospital with COVID-19 infection.
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http://dx.doi.org/10.1186/s12967-020-02524-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7491021PMC
September 2020

SOX9 promotes stress-responsive transcription of VGF nerve growth factor inducible gene in renal tubular epithelial cells.

J Biol Chem 2020 11 4;295(48):16328-16341. Epub 2020 Sep 4.

Division of Pharmaceutics and Pharmacology, College of Pharmacy and Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio, USA. Electronic address:

Acute kidney injury (AKI) is a common clinical condition associated with diverse etiologies and abrupt loss of renal function. In patients with sepsis, rhabdomyolysis, cancer, and cardiovascular disorders, the underlying disease or associated therapeutic interventions can cause hypoxia, cytotoxicity, and inflammatory insults to renal tubular epithelial cells (RTECs), resulting in the onset of AKI. To uncover stress-responsive disease-modifying genes, here we have carried out renal transcriptome profiling in three distinct murine models of AKI. We find that nerve growth factor inducible gene up-regulation is a common transcriptional stress response in RTECs to ischemia-, cisplatin-, and rhabdomyolysis-associated renal injury. The gene encodes a secretory peptide precursor protein that has critical neuroendocrine functions; however, its role in the kidneys remains unknown. Our functional studies show that RTEC-specific gene ablation exacerbates ischemia-, cisplatin-, and rhabdomyolysis-associated AKI and cisplatin-induced RTEC cell death Importantly, aggravation of cisplatin-induced renal injury caused by gene ablation is partly reversed by TLQP-21, a Vgf-derived peptide. Finally, and mechanistic studies showed that injury-induced up-regulation in RTECs is driven by the transcriptional regulator Sox9. These findings reveal a crucial downstream target of the Sox9-directed transcriptional program and identify as a stress-responsive protective gene in kidney tubular epithelial cells.
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http://dx.doi.org/10.1074/jbc.RA120.015110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7705303PMC
November 2020

SIOP PODC-adapted treatment guidelines for craniopharyngioma in low- and middle-income settings.

Pediatr Blood Cancer 2020 Aug 13:e28493. Epub 2020 Aug 13.

Department of Pediatric Oncology, Great North Children's Hospital, Newcastle upon Tyne, UK.

Pediatric craniopharyngioma is a rare tumor with excellent survival but significant long-term morbidities due to the loco-regional tumor growth or secondary to its treatment. Visual impairment, panhypopituitarism, hypothalamic damage, and behavioral changes are among the main challenges. This tumor should be managed under the care of a multidisciplinary team to determine the optimum treatment within the available resources. This is particularly important for low middle-income countries where resources are variable. This report provides risk-stratified management guidelines for children diagnosed with craniopharyngioma in a resource-limited setting.
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http://dx.doi.org/10.1002/pbc.28493DOI Listing
August 2020

ADAM10 mediates ectopic proximal tubule development and renal fibrosis through Notch signalling.

J Pathol 2020 11 24;252(3):274-289. Epub 2020 Sep 24.

Kidney Disease Center, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, PR China.

Disturbed intrauterine development increases the risk of renal disease. Various studies have reported that Notch signalling plays a significant role in kidney development and kidney diseases. A disintegrin and metalloproteinase domain 10 (ADAM10), an upstream protease of the Notch pathway, is also reportedly involved in renal fibrosis. However, how ADAM10 interacts with the Notch pathway and causes renal fibrosis is not fully understood. In this study, using a prenatal chlorpyrifos (CPF) exposure mouse model, we investigated the role of the ADAM10/Notch axis in kidney development and fibrosis. We found that prenatal CPF-exposure mice presented overexpression of Adam10, Notch1 and Notch2, and led to premature depletion of Six2 nephron progenitors and ectopic formation of proximal tubules (PTs) in the embryonic kidney. These abnormal phenotypic changes persisted in mature kidneys due to the continuous activation of ADAM10/Notch and showed aggravated renal fibrosis in adults. Finally, both ADAM10 and NOTCH2 expression were positively correlated with the degree of renal interstitial fibrosis in IgA nephropathy patients, and increased ADAM10 expression was negatively correlated with decreased kidney function evaluated by serum creatinine, cystatin C, and estimated glomerular filtration rate. Regression analysis also indicated that ADAM10 expression was an independent risk factor for fibrosis in IgAN. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.
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http://dx.doi.org/10.1002/path.5517DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7702158PMC
November 2020

Common Variants Coregulate Expression of GBA and Modifier Genes to Delay Parkinson's Disease Onset.

Mov Disord 2020 08 18;35(8):1346-1356. Epub 2020 Jun 18.

Liggins Institute, The University of Auckland, Auckland, New Zealand.

Background: GBA mutations are numerically the most significant genetic risk factor for Parkinson's disease (PD), yet these mutations have low penetrance, suggesting additional mechanisms.

Objectives: The objective of this study was to determine if the penetrance of GBA in PD can be explained by regulatory effects on GBA and modifier genes.

Methods: Genetic variants associated with the regulation of GBA were identified by screening 128 common single nucleotide polymorphisms (SNPs) in the GBA locus for spatial cis-expression quantitative trail locus (supported by chromatin interactions).

Results: We identified common noncoding SNPs within GBA that (1) regulate GBA expression in peripheral tissues, some of which display α-synuclein pathology and (2) coregulate potential modifier genes in the central nervous system and/or peripheral tissues. Haplotypes based on 3 of these SNPs delay disease onset by 5 years. In addition, SNPs on 6 separate chromosomes coregulate GBA expression specifically in either the substantia nigra or cortex, and their combined effect potentially modulates motor and cognitive symptoms, respectively.

Conclusions: This work provides a new perspective on the haplotype-specific effects of GBA and the genetic etiology of PD, expanding the role of GBA from the gene encoding the β-glucocerebrosidase (GCase) to that of a central regulator and modifier of PD onset, with GBA expression itself subject to distant regulation. Some idiopathic patients might possess insufficient GBA-encoded GCase activity in the substantia nigra as the result of distant regulatory variants and therefore might benefit from GBA-targeting therapeutics. The SNPs' regulatory impacts provide a plausible explanation for the variable phenotypes also observed in GBA-centric Gaucher's disease and dementia with Lewy bodies. © 2020 The Authors. Movement Disorders published by Wiley Periodicals, LLC on behalf of International Parkinson and Movement Disorder Society.
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http://dx.doi.org/10.1002/mds.28144DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7496525PMC
August 2020

AcrIF9 tethers non-sequence specific dsDNA to the CRISPR RNA-guided surveillance complex.

Nat Commun 2020 06 1;11(1):2730. Epub 2020 Jun 1.

Department of Microbiology and Immunology, Montana State University, 1156 South 11th Avenue, Bozeman, MT, 59717, USA.

Bacteria have evolved sophisticated adaptive immune systems, called CRISPR-Cas, that provide sequence-specific protection against phage infection. In turn, phages have evolved a broad spectrum of anti-CRISPRs that suppress these immune systems. Here we report structures of anti-CRISPR protein IF9 (AcrIF9) in complex with the type I-F CRISPR RNA-guided surveillance complex (Csy). In addition to sterically blocking the hybridization of complementary dsDNA to the CRISPR RNA, our results show that AcrIF9 binding also promotes non-sequence-specific engagement with dsDNA, potentially sequestering the complex from target DNA. These findings highlight the versatility of anti-CRISPR mechanisms utilized by phages to suppress CRISPR-mediated immune systems.
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http://dx.doi.org/10.1038/s41467-020-16512-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7264359PMC
June 2020

Clinicopathologic Characterization of Children With B-Cell Non-Hodgkin Lymphoma Over 10 Years at a Tertiary Center in Cape Town, South Africa.

J Pediatr Hematol Oncol 2020 05;42(4):e219-e227

Division of Haematology, National Health Laboratory Service (NHLS), Groote Schuur Hospital and the University of Cape Town.

Background: We characterized B-cell non-Hodgkin lymphoma (NHL) cases over 10 years at a tertiary children's hospital to contribute to the body of knowledge on pediatric lymphoma in developing countries with a high human immunodeficiency virus (HIV) burden.

Methods: A retrospective cohort study was carried out using clinical and laboratory records of children newly diagnosed with B-cell NHL from January 2005 to December 2014.

Results: Seventy-five children ≤15 years of age were included. The majority had Burkitt lymphoma (n=61). Overall, (n=19) were HIV positive and 16% (n=12) had concurrent active tuberculosis. Bulky disease was present in 65.7% (n=46) and 30.1% (n=22) were classified as Lymphomes Malins B risk group C. The 5-year survival estimates for HIV-negative and HIV-positive children were similar in our cohort: 81% versus 79% for event-free survival and 85% versus 83.9% for overall survival. Of 3 children with Burkitt lymphoma, HIV, and Lymphomes Malins B group C, 2 died within 1 year.

Conclusions: Irrespective of HIV status, the survival of children in our B-cell NHL cohort compares favorably with cure rates in developed nations, although advanced disease remains associated with a poor prognosis. Characterization of childhood NHL cases contributes to accurate risk stratification and tailored treatment.
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http://dx.doi.org/10.1097/MPH.0000000000001709DOI Listing
May 2020

Listeria Phages Induce Cas9 Degradation to Protect Lysogenic Genomes.

Cell Host Microbe 2020 07 22;28(1):31-40.e9. Epub 2020 Apr 22.

Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Innovative Genomics Institute, Berkeley, CA 94720, USA. Electronic address:

Bacterial CRISPR-Cas systems employ RNA-guided nucleases to destroy phage (viral) DNA. Phages, in turn, have evolved diverse "anti-CRISPR" proteins (Acrs) to counteract acquired immunity. In Listeria monocytogenes, prophages encode two to three distinct anti-Cas9 proteins, with acrIIA1 always present. However, the significance of AcrIIA1's pervasiveness and its mechanism are unknown. Here, we report that AcrIIA1 binds with high affinity to Cas9 via the catalytic HNH domain. During lysogeny in Listeria, AcrIIA1 triggers Cas9 degradation. During lytic infection, however, AcrIIA1 fails to block Cas9 due to its multi-step inactivation mechanism. Thus, phages encode an additional Acr that rapidly binds and inactivates Cas9. AcrIIA1 also uniquely inhibits a highly diverged Cas9 found in Listeria (similar to SauCas9) and Type II-C Cas9s, likely due to Cas9 HNH domain conservation. In summary, Listeria phages inactivate Cas9 in lytic growth using variable, narrow-spectrum inhibitors, while the broad-spectrum AcrIIA1 stimulates Cas9 degradation for protection of the lysogenic genome.
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http://dx.doi.org/10.1016/j.chom.2020.04.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7351598PMC
July 2020

The Utility of Human Kidney Organoids in Modeling Kidney Disease.

Semin Nephrol 2020 03;40(2):188-198

Department of Molecular Medicine and Pathology, School of Medical Sciences, University of Auckland, Auckland, New Zealand.

The formation of three-dimensional kidney tissue (organoids) from human pluripotent stem cell lines provides a valuable tool to examine kidney function in an in vitro model and could be used for regenerative medicine approaches. Kidney organoids have the potential to model kidney diseases and congenital defects, be used for drug development, and to further our understanding of acute kidney injury, fibrosis, and chronic kidney disease. In this review, we examine the current stage of pluripotent stem cell-derived kidney organoid technology, challenges, shortcomings, and regenerative potential of kidney organoids in the future.
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http://dx.doi.org/10.1016/j.semnephrol.2020.01.009DOI Listing
March 2020

Use of Human Induced Pluripotent Stem Cells and Kidney Organoids To Develop a Cysteamine/mTOR Inhibition Combination Therapy for Cystinosis.

J Am Soc Nephrol 2020 05 20;31(5):962-982. Epub 2020 Mar 20.

Department of Molecular Medicine and Pathology, The University of Auckland, Auckland, New Zealand

Background: Mutations in -a gene encoding the cystine transporter cystinosin-cause the rare, autosomal, recessive, lysosomal-storage disease cystinosis. Research has also implicated cystinosin in modulating the mTORC1 pathway, which serves as a core regulator of cellular metabolism, proliferation, survival, and autophagy. In its severest form, cystinosis is characterized by cystine accumulation, renal proximal tubule dysfunction, and kidney failure. Because treatment with the cystine-depleting drug cysteamine only slows disease progression, there is an urgent need for better treatments.

Methods: To address a lack of good human-based cell culture models for studying cystinosis, we generated the first human induced pluripotent stem cell (iPSC) and kidney organoid models of the disorder. We used a variety of techniques to examine hallmarks of cystinosis-including cystine accumulation, lysosome size, the autophagy pathway, and apoptosis-and performed RNA sequencing on isogenic lines to identify differentially expressed genes in the cystinosis models compared with controls.

Results: Compared with controls, these cystinosis models exhibit elevated cystine levels, increased apoptosis, and defective basal autophagy. Cysteamine treatment ameliorates this phenotype, except for abnormalities in apoptosis and basal autophagy. We found that treatment with everolimus, an inhibitor of the mTOR pathway, reduces the number of large lysosomes, decreases apoptosis, and activates autophagy, but it does not rescue the defect in cystine loading. However, dual treatment of cystinotic iPSCs or kidney organoids with cysteamine and everolimus corrects all of the observed phenotypic abnormalities.

Conclusions: These observations suggest that combination therapy with a cystine-depleting drug such as cysteamine and an mTOR pathway inhibitor such as everolimus has potential to improve treatment of cystinosis.
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http://dx.doi.org/10.1681/ASN.2019070712DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7217405PMC
May 2020

Anti-CRISPRs: Protein Inhibitors of CRISPR-Cas Systems.

Annu Rev Biochem 2020 06 18;89:309-332. Epub 2020 Mar 18.

RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA; email:

Clustered regularly interspaced short palindromic repeats (CRISPR) together with their accompanying (CRISPR-associated) genes are found frequently in bacteria and archaea, serving to defend against invading foreign DNA, such as viral genomes. CRISPR-Cas systems provide a uniquely powerful defense because they can adapt to newly encountered genomes. The adaptive ability of these systems has been exploited, leading to their development as highly effective tools for genome editing. The widespread use of CRISPR-Cas systems has driven a need for methods to control their activity. This review focuses on anti-CRISPRs (Acrs), proteins produced by viruses and other mobile genetic elements that can potently inhibit CRISPR-Cas systems. Discovered in 2013, there are now 54 distinct families of these proteins described, and the functional mechanisms of more than a dozen have been characterized in molecular detail. The investigation of Acrs is leading to a variety of practical applications and is providing exciting new insight into the biology of CRISPR-Cas systems.
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http://dx.doi.org/10.1146/annurev-biochem-011420-111224DOI Listing
June 2020

Evaluation of cisplatin-induced injury in human kidney organoids.

Am J Physiol Renal Physiol 2020 04 9;318(4):F971-F978. Epub 2020 Mar 9.

Department of Molecular Medicine and Pathology, University of Auckland, Auckland, New Zealand.

Acute kidney injury (AKI) remains a major global healthcare problem, and there is a need to develop human-based models to study AKI in vitro. Toward this goal, we have characterized induced pluripotent stem cell-derived human kidney organoids and their response to cisplatin, a chemotherapeutic drug that induces AKI and preferentially damages the proximal tubule. We found that a single treatment with 50 µM cisplatin induces hepatitis A virus cellular receptor 1 () and C-X-C motif chemokine ligand 8 () expression, DNA damage (γH2AX), and cell death in the organoids but greatly impairs organoid viability. DNA damage was not specific to the proximal tubule but also affected the distal tubule and interstitial cell populations. This lack of specificity correlated with low expression of proximal tubule-specific organic cation transporter 2 () for cisplatin. To improve viability, we developed a repeated low-dose regimen of 4 × 5 µM cisplatin over 7 days and found this caused less toxicity while still inducing a robust injury response that included secretion of known AKI biomarkers and inflammatory cytokines. This work validates the use of human kidney organoids to model aspects of cisplatin-induced injury, with the potential to identify new AKI biomarkers and develop better therapies.
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http://dx.doi.org/10.1152/ajprenal.00597.2019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395477PMC
April 2020

Global Retinoblastoma Presentation and Analysis by National Income Level.

JAMA Oncol 2020 05;6(5):685-695

Imam Hussein Cancer Center, Karbala, Iraq.

Importance: Early diagnosis of retinoblastoma, the most common intraocular cancer, can save both a child's life and vision. However, anecdotal evidence suggests that many children across the world are diagnosed late. To our knowledge, the clinical presentation of retinoblastoma has never been assessed on a global scale.

Objectives: To report the retinoblastoma stage at diagnosis in patients across the world during a single year, to investigate associations between clinical variables and national income level, and to investigate risk factors for advanced disease at diagnosis.

Design, Setting, And Participants: A total of 278 retinoblastoma treatment centers were recruited from June 2017 through December 2018 to participate in a cross-sectional analysis of treatment-naive patients with retinoblastoma who were diagnosed in 2017.

Main Outcomes And Measures: Age at presentation, proportion of familial history of retinoblastoma, and tumor stage and metastasis.

Results: The cohort included 4351 new patients from 153 countries; the median age at diagnosis was 30.5 (interquartile range, 18.3-45.9) months, and 1976 patients (45.4%) were female. Most patients (n = 3685 [84.7%]) were from low- and middle-income countries (LMICs). Globally, the most common indication for referral was leukocoria (n = 2638 [62.8%]), followed by strabismus (n = 429 [10.2%]) and proptosis (n = 309 [7.4%]). Patients from high-income countries (HICs) were diagnosed at a median age of 14.1 months, with 656 of 666 (98.5%) patients having intraocular retinoblastoma and 2 (0.3%) having metastasis. Patients from low-income countries were diagnosed at a median age of 30.5 months, with 256 of 521 (49.1%) having extraocular retinoblastoma and 94 of 498 (18.9%) having metastasis. Lower national income level was associated with older presentation age, higher proportion of locally advanced disease and distant metastasis, and smaller proportion of familial history of retinoblastoma. Advanced disease at diagnosis was more common in LMICs even after adjusting for age (odds ratio for low-income countries vs upper-middle-income countries and HICs, 17.92 [95% CI, 12.94-24.80], and for lower-middle-income countries vs upper-middle-income countries and HICs, 5.74 [95% CI, 4.30-7.68]).

Conclusions And Relevance: This study is estimated to have included more than half of all new retinoblastoma cases worldwide in 2017. Children from LMICs, where the main global retinoblastoma burden lies, presented at an older age with more advanced disease and demonstrated a smaller proportion of familial history of retinoblastoma, likely because many do not reach a childbearing age. Given that retinoblastoma is curable, these data are concerning and mandate intervention at national and international levels. Further studies are needed to investigate factors, other than age at presentation, that may be associated with advanced disease in LMICs.
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http://dx.doi.org/10.1001/jamaoncol.2019.6716DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7047856PMC
May 2020

SIOP PODC adapted risk stratification and treatment guidelines: Recommendations for acute myeloid leukemia in resource-limited settings.

Pediatr Blood Cancer 2019 Nov 27:e28087. Epub 2019 Nov 27.

Department of Pediatric Oncology, Emma Children's Hospital, Amsterdam UMC, Vrije Universiteit, Amsterdam, The Netherlands.

In low- and middle-income countries (LMICs), limited resources, suboptimal risk stratification, and disproportionate patient-to-infrastructure ratio result in low survival of patients with acute myeloid leukemia (AML). A high incidence of relapse, inherent to the biology, renders management arduous. The challenge of treating AML in LMICs is of balancing the intensity of myelosuppressive chemotherapy, which appears necessary for cure, with available supportive care, which influences treatment-related mortality. The recommendations outlined in this paper are based on published evidence and expert opinion. The principle of this adapted protocol is to tailor treatment to available resources, reduce preventable toxic death, and direct limited resources toward those children who are most likely to be cured.
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http://dx.doi.org/10.1002/pbc.28087DOI Listing
November 2019

Anti-CRISPR AcrIIA5 Potently Inhibits All Cas9 Homologs Used for Genome Editing.

Cell Rep 2019 11;29(7):1739-1746.e5

Department of Molecular Genetics, University of Toronto, Toronto, ON M5G 1M1, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5G 1M1, Canada. Electronic address:

CRISPR-Cas9 systems provide powerful tools for genome editing. However, optimal employment of this technology will require control of Cas9 activity so that the timing, tissue specificity, and accuracy of editing may be precisely modulated. Anti-CRISPR proteins, which are small, naturally occurring inhibitors of CRISPR-Cas systems, are well suited for this purpose. A number of anti-CRISPR proteins have been shown to potently inhibit subgroups of CRISPR-Cas9 systems, but their maximal inhibitory activity is generally restricted to specific Cas9 homologs. Since Cas9 homologs vary in important properties, differing Cas9s may be optimal for particular genome-editing applications. To facilitate the practical exploitation of multiple Cas9 homologs, here we identify one anti-CRISPR, called AcrIIA5, that potently inhibits nine diverse type II-A and type II-C Cas9 homologs, including those currently used for genome editing. We show that the activity of AcrIIA5 results in partial in vivo cleavage of a single-guide RNA (sgRNA), suggesting that its mechanism involves RNA interaction.
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http://dx.doi.org/10.1016/j.celrep.2019.10.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6910239PMC
November 2019

Anti-CRISPR-Associated Proteins Are Crucial Repressors of Anti-CRISPR Transcription.

Cell 2019 09 29;178(6):1452-1464.e13. Epub 2019 Aug 29.

Department of Molecular Genetics, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Biochemistry, University of Toronto, Toronto, ON M5S 1A8, Canada. Electronic address:

Phages express anti-CRISPR (Acr) proteins to inhibit CRISPR-Cas systems that would otherwise destroy their genomes. Most acr genes are located adjacent to anti-CRISPR-associated (aca) genes, which encode proteins with a helix-turn-helix DNA-binding motif. The conservation of aca genes has served as a signpost for the identification of acr genes, but the function of the proteins encoded by these genes has not been investigated. Here we reveal that an acr-associated promoter drives high levels of acr transcription immediately after phage DNA injection and that Aca proteins subsequently repress this transcription. Without Aca activity, this strong transcription is lethal to a phage. Our results demonstrate how sufficient levels of Acr proteins accumulate early in the infection process to inhibit existing CRISPR-Cas complexes in the host cell. They also imply that the conserved role of Aca proteins is to mitigate the deleterious effects of strong constitutive transcription from acr promoters.
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http://dx.doi.org/10.1016/j.cell.2019.07.046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6754177PMC
September 2019

Inhibition of CRISPR-Cas9 ribonucleoprotein complex assembly by anti-CRISPR AcrIIC2.

Nat Commun 2019 06 26;10(1):2806. Epub 2019 Jun 26.

Department of Biochemistry, University of Toronto, 661 University Avenue, Suite 1600, Toronto, ON, M5G 1M1, Canada.

CRISPR-Cas adaptive immune systems function to protect bacteria from invasion by foreign genetic elements. The CRISPR-Cas9 system has been widely adopted as a powerful genome-editing tool, and phage-encoded inhibitors, known as anti-CRISPRs, offer a means of regulating its activity. Here, we report the crystal structures of anti-CRISPR protein AcrIIC2 alone and in complex with Nme1Cas9. We demonstrate that AcrIIC2 inhibits Cas9 through interactions with the positively charged bridge helix, thereby preventing sgRNA loading. In vivo phage plaque assays and in vitro DNA cleavage assays show that AcrIIC2 mediates its activity through a large electronegative surface. This work shows that anti-CRISPR activity can be mediated through the inhibition of Cas9 complex assembly.
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http://dx.doi.org/10.1038/s41467-019-10577-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6594998PMC
June 2019

Phage tail fibre assembly proteins employ a modular structure to drive the correct folding of diverse fibres.

Nat Microbiol 2019 10 17;4(10):1645-1653. Epub 2019 Jun 17.

Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada.

Phage tail fibres are elongated protein assemblies capable of specific recognition of bacterial surfaces during the first step of viral infection. The folding of these complex trimeric structures often requires a phage-encoded tail fibre assembly (Tfa) protein. Despite the wide occurrence of Tfa proteins, their functional mechanism has not been elucidated. Here, we investigate the tail fibre and Tfa of Escherichia coli phage Mu. We demonstrate that Tfa forms a stable complex with the tail fibre, and present a 2.1 Å resolution X-ray crystal structure of this complex. We find that Tfa proteins are comprised of two domains: a non-conserved N-terminal domain that binds to the C-terminal region of the fibre and a conserved C-terminal domain that probably mediates fibre oligomerization and assembly. Tfa forms rapidly exchanging multimers on its own, but not a stable trimer, implying that Tfa does not specify the trimeric state of the fibre. We propose that the key conserved role of Tfa is to ensure that fibre assembly and multimerization initiates at the C terminus, ensuring that the intertwined and repetitive structural elements of fibres come together in the correct sequence. The universal importance of correctly aligning the C termini of phage fibres is highlighted by our work.
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http://dx.doi.org/10.1038/s41564-019-0477-7DOI Listing
October 2019

Transcriptional profiling of the zebrafish proximal tubule.

Am J Physiol Renal Physiol 2019 08 12;317(2):F478-F488. Epub 2019 Jun 12.

Department of Molecular Medicine and Pathology, University of Auckland, Auckland, New Zealand.

The hepatocyte nuclear factor-1β (Hnf1b) transcription factor is a key regulator of kidney tubule formation and is associated with a syndrome of renal cysts and early onset diabetes. To further our understanding of Hnf1b in the developing zebrafish kidney, we performed RNA sequencing analysis of proximal tubules from -deficient larvae. This analysis revealed an enrichment of gene transcripts encoding transporters of the solute carrier (SLC) superfamily, including multiple members of and glucose transporters. An investigation of expression of , and as well as a poorly studied glucose/mannose transporter encoded by revealed that these genes undergo dynamic spatiotemporal changes during tubule formation and maturation. A comparative analysis of zebrafish SLC genes with those expressed in mouse proximal tubules showed a substantial overlap at the level of gene families, indicating a high degree of functional conservation between zebrafish and mammalian proximal tubules. Taken together, our findings are consistent with a role for Hnf1b as a critical determinant of proximal tubule transport function by acting upstream of a large number of SLC genes and validate the zebrafish as a physiologically relevant model of the mammalian proximal tubule.
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http://dx.doi.org/10.1152/ajprenal.00174.2019DOI Listing
August 2019