Publications by authors named "Joseph M Replogle"

19 Publications

  • Page 1 of 1

Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing.

Cell 2021 Apr 9;184(9):2503-2519.e17. Epub 2021 Apr 9.

Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, CA 94158, USA; Whitehead Institute for Biomedical Research, Massachusetts Institute of Technology, Cambridge 02142, USA. Electronic address:

A general approach for heritably altering gene expression has the potential to enable many discovery and therapeutic efforts. Here, we present CRISPRoff-a programmable epigenetic memory writer consisting of a single dead Cas9 fusion protein that establishes DNA methylation and repressive histone modifications. Transient CRISPRoff expression initiates highly specific DNA methylation and gene repression that is maintained through cell division and differentiation of stem cells to neurons. Pairing CRISPRoff with genome-wide screens and analysis of chromatin marks establishes rules for heritable gene silencing. We identify single guide RNAs (sgRNAs) capable of silencing the large majority of genes including those lacking canonical CpG islands (CGIs) and reveal a wide targeting window extending beyond annotated CGIs. The broad ability of CRISPRoff to initiate heritable gene silencing even outside of CGIs expands the canonical model of methylation-based silencing and enables diverse applications including genome-wide screens, multiplexed cell engineering, enhancer silencing, and mechanistic exploration of epigenetic inheritance.
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http://dx.doi.org/10.1016/j.cell.2021.03.025DOI Listing
April 2021

GIGYF2 and 4EHP Inhibit Translation Initiation of Defective Messenger RNAs to Assist Ribosome-Associated Quality Control.

Mol Cell 2020 09 28;79(6):950-962.e6. Epub 2020 Jul 28.

Department of Embryology, Carnegie Institution for Science, Baltimore, MD 21218, USA. Electronic address:

Ribosome-associated quality control (RQC) pathways protect cells from toxicity caused by incomplete protein products resulting from translation of damaged or problematic mRNAs. Extensive work in yeast has identified highly conserved mechanisms that lead to degradation of faulty mRNA and partially synthesized polypeptides. Here we used CRISPR-Cas9-based screening to search for additional RQC strategies in mammals. We found that failed translation leads to specific inhibition of translation initiation on that message. This negative feedback loop is mediated by two translation inhibitors, GIGYF2 and 4EHP. Model substrates and growth-based assays established that inhibition of additional rounds of translation acts in concert with known RQC pathways to prevent buildup of toxic proteins. Inability to block translation of faulty mRNAs and subsequent accumulation of partially synthesized polypeptides could explain the neurodevelopmental and neuropsychiatric disorders observed in mice and humans with compromised GIGYF2 function.
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http://dx.doi.org/10.1016/j.molcel.2020.07.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7891188PMC
September 2020

Combinatorial single-cell CRISPR screens by direct guide RNA capture and targeted sequencing.

Nat Biotechnol 2020 08 30;38(8):954-961. Epub 2020 Mar 30.

Lewis-Sigler Institute for Integrative Genomics, Princeton University, Princeton, NJ, USA.

Single-cell CRISPR screens enable the exploration of mammalian gene function and genetic regulatory networks. However, use of this technology has been limited by reliance on indirect indexing of single-guide RNAs (sgRNAs). Here we present direct-capture Perturb-seq, a versatile screening approach in which expressed sgRNAs are sequenced alongside single-cell transcriptomes. Direct-capture Perturb-seq enables detection of multiple distinct sgRNA sequences from individual cells and thus allows pooled single-cell CRISPR screens to be easily paired with combinatorial perturbation libraries that contain dual-guide expression vectors. We demonstrate the utility of this approach for high-throughput investigations of genetic interactions and, leveraging this ability, dissect epistatic interactions between cholesterol biogenesis and DNA repair. Using direct capture Perturb-seq, we also show that targeting individual genes with multiple sgRNAs per cell improves efficacy of CRISPR interference and activation, facilitating the use of compact, highly active CRISPR libraries for single-cell screens. Last, we show that hybridization-based target enrichment permits sensitive, specific sequencing of informative transcripts from single-cell RNA-seq experiments.
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http://dx.doi.org/10.1038/s41587-020-0470-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7416462PMC
August 2020

Exploring genetic interaction manifolds constructed from rich single-cell phenotypes.

Science 2019 08 8;365(6455):786-793. Epub 2019 Aug 8.

Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94158, USA.

How cellular and organismal complexity emerges from combinatorial expression of genes is a central question in biology. High-content phenotyping approaches such as Perturb-seq (single-cell RNA-sequencing pooled CRISPR screens) present an opportunity for exploring such genetic interactions (GIs) at scale. Here, we present an analytical framework for interpreting high-dimensional landscapes of cell states (manifolds) constructed from transcriptional phenotypes. We applied this approach to Perturb-seq profiling of strong GIs mined from a growth-based, gain-of-function GI map. Exploration of this manifold enabled ordering of regulatory pathways, principled classification of GIs (e.g., identifying suppressors), and mechanistic elucidation of synergistic interactions, including an unexpected synergy between and driving erythroid differentiation. Finally, we applied recommender system machine learning to predict interactions, facilitating exploration of vastly larger GI manifolds.
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http://dx.doi.org/10.1126/science.aax4438DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6746554PMC
August 2019

A human microglia-like cellular model for assessing the effects of neurodegenerative disease gene variants.

Sci Transl Med 2017 Dec;9(421)

Program in Medical and Population Genetics, Broad Institute, 7 Cambridge Center, Cambridge, MA 02142, USA.

Microglia are emerging as a key cell type in neurodegenerative diseases, yet human microglia are challenging to study in vitro. We developed an in vitro cell model system composed of human monocyte-derived microglia-like (MDMi) cells that recapitulated key aspects of microglia phenotype and function. We then used this model system to perform an expression quantitative trait locus (eQTL) study examining 94 genes from loci associated with Alzheimer's disease, Parkinson's disease, and multiple sclerosis. We found six loci (, , , , , and ) in which the risk haplotype drives the association with both disease susceptibility and altered expression of a nearby gene (cis-eQTL). In the and loci, the cis-eQTL was found in the MDMi cells but not in human peripheral blood monocytes, suggesting that differentiation of monocytes into microglia-like cells led to the acquisition of a cellular state that could reveal the functional consequences of certain genetic variants. We further validated the effect of risk haplotypes at the protein level for and , and we confirmed that the risk haplotype altered phagocytosis by the MDMi cells. We propose that increased gene expression by MDMi cells could be a functional outcome of , a single-nucleotide polymorphism in the locus that is associated with Parkinson's disease.
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http://dx.doi.org/10.1126/scitranslmed.aai7635DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5945290PMC
December 2017

Genetic architecture of age-related cognitive decline in African Americans.

Neurol Genet 2017 Feb 21;3(1):e125. Epub 2016 Dec 21.

Program in Translational NeuroPsychiatric Genomics (T.R., L.B.C., J.M.R., P.L.D.J.), Institute for the Neurosciences, Departments of Neurology and Psychiatry, Center for Neurologic Disease (T.R., A.W., A.K., P.L.D.J.), Department of Neurology, and Division of Genetics (T.R., L.B.C., P.L.D.J.), Department of Medicine, Brigham and Women's Hospital, Boston, MA; Harvard Medical School (T.R., L.B.C., P.L.D.J.), Boston, MA; Program in Medical and Population genetics (T.R., L.B.C., C.M., J.M.R., P.L.D.J.), The Broad Institute, Cambridge, MA; Section of Genetic Medicine (B.S.), Department of Medicine, and Institute for Genomics and Systems Biology (B.S.), University of Chicago, IL; Indiana University Center for Aging Research (H.C.H.); Department of Psychiatry (F.W.U., H.C.H., K.S.H.), Department of Biostatistics (S.G.), Indiana University School of Medicine; Department of Medical and Molecular Genetics (J.M., T.F.), Indiana University, Indianapolis; Rush Institute for Healthy Aging (D.A.V.), Department of Internal Medicine, Department of Neurology (L.B., D.A.B.), and Rush Alzheimer's Disease Center (L.Y., L.B., D.A.B.), Rush University Medical Center, Chicago, IL. T.R. is currently affiliated with Ronald M. Loeb Center for Alzheimer's Disease, Departments of Neuroscience, and Genetics and Genomic Sciences, Mount Sinai School of Medicine, New York.

Objective: To identify genetic risk factors associated with susceptibility to age-related cognitive decline in African Americans (AAs).

Methods: We performed a genome-wide association study (GWAS) and an admixture-mapping scan in 3,964 older AAs from 5 longitudinal cohorts; for each participant, we calculated a slope of an individual's global cognitive change from neuropsychological evaluations. We also performed a pathway-based analysis of the age-related cognitive decline GWAS.

Results: We found no evidence to support the existence of a genomic region which has a strongly different contribution to age-related cognitive decline in African and European genomes. Known Alzheimer disease (AD) susceptibility variants in the and loci do influence this trait in AAs. Of interest, our pathway-based analyses returned statistically significant results highlighting a shared risk from lipid/metabolism and protein tyrosine signaling pathways between cognitive decline and AD, but the role of inflammatory pathways is polarized, being limited to AD susceptibility.

Conclusions: The genetic architecture of aging-related cognitive in AA individuals is largely similar to that of individuals of European descent. In both populations, we note a surprising lack of enrichment for immune pathways in the genetic risk for cognitive decline, despite strong enrichment of these pathways among genetic risk factors for AD.
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http://dx.doi.org/10.1212/NXG.0000000000000125DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5206965PMC
February 2017

Trans-pQTL study identifies immune crosstalk between Parkinson and Alzheimer loci.

Neurol Genet 2016 Aug 26;2(4):e90. Epub 2016 Jul 26.

Ann Romney Center for Neurologic Diseases (G.C., C.C.W., P.A.W., M.C., J.M.R., L.R.G., N.E.C., K.J.R., L.B.C., P.L.D.J., E.M.B.), Program in Translational NeuroPsychiatric Genomics (G.C., C.C.W., P.A.W., M.C., J.M.R., L.R.G., N.E.C., K.J.R., L.B.C., P.L.D.J., E.M.B.), Institute for the Neurosciences, Departments of Neurology and Psychiatry, Center for Alzheimer's Research and Treatment (K.A.J., R.A.S.), Department of Neurology, Brigham and Women's Hospital, Boston, MA; Program in Medical and Population Genetics (G.C., C.C.W., P.A.W., M.C., J.M.R., L.R.G., N.E.C., K.J.R., L.B.C., P.L.D.J., E.M.B.), Broad Institute, Cambridge, MA; Harvard Medical School (G.C., J.M.R., K.J.R., K.A.J., L.B.C., R.A.S., P.L.D.J., E.M.B.), Boston, MA; Department of Neurology (K.A.J., R.A.S.), Massachusetts General Hospital, Boston; and Rush Alzheimer's Disease Center (J.A.S., D.A.B.), Rush University Medical Center, Chicago, IL.

Objective: Given evidence from genetic studies, we hypothesized that there may be a shared component to the role of myeloid function in Parkinson and Alzheimer disease (PD and AD) and assessed whether PD susceptibility variants influenced protein expression of well-established AD-associated myeloid genes in human monocytes.

Methods: We repurposed data in which AD-related myeloid proteins CD33, TREM1, TREM2, TREML2, TYROBP, and PTK2B were measured by flow cytometry in monocytes from 176 participants of the PhenoGenetic Project (PGP) and Harvard Aging Brain Study. Linear regression was used to identify associations between 24 PD risk variants and protein expression. The 2 cohorts were meta-analyzed in a discovery analysis, and the 4 most strongly suggestive results were validated in an independent cohort of 50 PGP participants.

Results: We discovered and validated an association between the PD risk allele rs12456492(G) in the RIT2 locus and increased CD33 expression (p joint = 3.50 × 10(-5)) and found strongly suggestive evidence that rs11060180(A) in the CCDC62/HIP1R locus decreased PTK2B expression (p joint = 1.12 × 10(-4)). Furthermore, in older individuals, increased CD33 expression on peripheral monocytes was associated with a greater burden of parkinsonism (p = 0.047), particularly bradykinesia (p = 6.64 × 10(-3)).

Conclusions: We find that the rs12456492 PD risk variant affects expression of AD-associated protein CD33 in peripheral monocytes, which suggests that genetic factors for these 2 diseases may converge to influence overlapping innate immune-mediated mechanisms that contribute to neurodegeneration. Furthermore, the effect of the rs12456492(G) PD risk allele on increased CD33 suggests that the inhibition of certain myeloid functions may contribute to PD susceptibility, as is the case for AD.
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http://dx.doi.org/10.1212/NXG.0000000000000090DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4962525PMC
August 2016

Nucleosomes impede Cas9 access to DNA in vivo and in vitro.

Elife 2016 03 17;5. Epub 2016 Mar 17.

Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, United States.

The prokaryotic CRISPR (clustered regularly interspaced palindromic repeats)-associated protein, Cas9, has been widely adopted as a tool for editing, imaging, and regulating eukaryotic genomes. However, our understanding of how to select single-guide RNAs (sgRNAs) that mediate efficient Cas9 activity is incomplete, as we lack insight into how chromatin impacts Cas9 targeting. To address this gap, we analyzed large-scale genetic screens performed in human cell lines using either nuclease-active or nuclease-dead Cas9 (dCas9). We observed that highly active sgRNAs for Cas9 and dCas9 were found almost exclusively in regions of low nucleosome occupancy. In vitro experiments demonstrated that nucleosomes in fact directly impede Cas9 binding and cleavage, while chromatin remodeling can restore Cas9 access. Our results reveal a critical role of eukaryotic chromatin in dictating the targeting specificity of this transplanted bacterial enzyme, and provide rules for selecting Cas9 target sites distinct from and complementary to those based on sequence properties.
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http://dx.doi.org/10.7554/eLife.12677DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4861601PMC
March 2016

Rheumatoid arthritis-associated RBPJ polymorphism alters memory CD4+ T cells.

Hum Mol Genet 2016 Jan 24;25(2):404-17. Epub 2015 Nov 24.

Ann Romney Center for Neurologic Diseases, Program in Translational NeuroPsychiatric Genomics, Department of Neurology, Brigham and Women's Hospital and Harvard Medical School, Broad Institute at Harvard University and MIT, NRB-641, 77 Avenue Louis Pasteur, Boston, MA 02115, USA and

Notch signaling has recently emerged as an important regulator of immune responses in autoimmune diseases. The recombination signal-binding protein for immunoglobulin kappa J region (RBPJ) is a transcriptional repressor, but converts into a transcriptional activator upon activation of the canonical Notch pathway. Genome-wide association studies of rheumatoid arthritis (RA) identified a susceptibility locus, rs874040(CC), which implicated the RBPJ gene. Here, chromatin state mapping generated using the chromHMM algorithm reveals strong enhancer regions containing DNase I hypersensitive sites overlapping the rs874040 linkage disequilibrium block in human memory, but not in naïve CD4(+) T cells. The rs874040 overlapping this chromatin state was associated with increased RBPJ expression in stimulated memory CD4(+) T cells from healthy subjects homozygous for the risk allele (CC) compared with memory CD4(+) T cells bearing the protective allele (GG). Transcriptomic analysis of rs874040(CC) memory T cells showed a repression of canonical Notch target genes IL (interleukin)-9, IL-17 and interferon (IFN)γ in the basal state. Interestingly, activation of the Notch pathway using soluble Notch ligand, Jagged2-Fc, induced IL-9 and IL-17A while delta-like 4Fc, another Notch ligand, induced higher IFNγ expression in the rs874040(CC) memory CD4(+) T cells compared with their rs874040(GG) counterparts. In RA, RBPJ expression is elevated in memory T cells from RA patients compared with control subjects, and this was associated with induced inflammatory cytokines IL-9, IL-17A and IFNγ in response to Notch ligation in vitro. These findings demonstrate that the rs874040(CC) allele skews memory T cells toward a pro-inflammatory phenotype involving Notch signaling, thus increasing the susceptibility to develop RA.
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http://dx.doi.org/10.1093/hmg/ddv474DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4881829PMC
January 2016

CD33 modulates TREM2: convergence of Alzheimer loci.

Nat Neurosci 2015 Nov 28;18(11):1556-8. Epub 2015 Sep 28.

Ann Romney Center for Neurologic Diseases, Brigham and Women's Hospital, Boston, Massachusetts, USA.

We used a protein quantitative trait analysis in monocytes from 226 individuals to evaluate cross-talk between Alzheimer loci. The NME8 locus influenced PTK2B and the CD33 risk allele led to greater TREM2 expression. There was also a decreased TREM1/TREM2 ratio with a TREM1 risk allele, decreased TREM2 expression with CD33 suppression and elevated cortical TREM2 mRNA expression with amyloid pathology.
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http://dx.doi.org/10.1038/nn.4126DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4682915PMC
November 2015

Reply: To PMID 25545807.

Ann Neurol 2015 Oct 22;78(4):659-60. Epub 2015 Aug 22.

Program in Translational NeuroPsychiatric Genomics, Institute for the Neurosciences, Departments of Neurology and Psychiatry, Brigham and Women's Hospital, Boston, MA.

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http://dx.doi.org/10.1002/ana.24456DOI Listing
October 2015

A pharmacogenetic study implicates SLC9a9 in multiple sclerosis disease activity.

Ann Neurol 2015 Jul 14;78(1):115-27. Epub 2015 May 14.

Ann Romney Center for Neurologic Diseases, Department of Neurology, Brigham and Women's Hospital, Boston, MA.

Objective: A proportion of multiple sclerosis (MS) patients experience disease activity despite treatment. The early identification of the most effective drug is critical to impact long-term outcome and to move toward a personalized approach. The aim of the present study is to identify biomarkers for further clinical development and to yield insights into the pathophysiology of disease activity.

Methods: We performed a genome-wide association study in interferon-β (IFNβ)-treated MS patients followed by validation in 3 independent cohorts. The role of the validated variant was examined in several RNA data sets, and the function of the presumed target gene was explored using an RNA interference approach in primary T cells in vitro.

Results: We found an association between rs9828519(G) and nonresponse to IFNβ (pdiscovery = 4.43 × 10(-8)) and confirmed it in a meta-analysis across 3 replication data sets (preplication = 7.78 × 10(-4)). Only 1 gene is found in the linkage disequilibrium block containing rs9828519: SLC9A9. Exploring the function of this gene, we see that SLC9A9 mRNA expression is diminished in MS subjects who are more likely to have relapses. Moreover, SLC9A9 knockdown in T cells in vitro leads an increase in expression of IFNγ, which is a proinflammatory molecule.

Interpretation: This study identifies and validates the role of rs9828519, an intronic variant in SLC9A9, in IFNβ-treated subjects, demonstrating a successful pharmacogenetic screen in MS. Functional characterization suggests that SLC9A9, an Na(+) -H(+) exchanger found in endosomes, appears to influence the differentiation of T cells to a proinflammatory fate and may have a broader role in MS disease activity, outside of IFNβ treatment.
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http://dx.doi.org/10.1002/ana.24429DOI Listing
July 2015

A TREM1 variant alters the accumulation of Alzheimer-related amyloid pathology.

Ann Neurol 2015 Mar;77(3):469-77

Program in Translational NeuroPsychiatric Genomics, Institute for the Neurosciences, Departments of Neurology and Psychiatry, Brigham and Women's Hospital, Boston, MA; Center for Neurologic Diseases, Brigham and Women's Hospital, Boston, MA; Program in Medical and Population Genetics, Broad Institute, Cambridge, MA; Harvard Medical School, Boston, MA.

Objective: Genome-wide association studies have linked variants in TREM2 (triggering receptor expressed on myeloid cells 2) and TREML2 with Alzheimer disease (AD) and AD endophenotypes. Here, we pursue a targeted analysis of the TREM locus in relation to cognitive decline and pathological features of AD.

Methods: Clinical, cognitive, and neuropathological phenotypes were collected in 3 prospective cohorts on aging (n = 3,421 subjects). Our primary analysis was an association with neuritic plaque pathology. To functionally characterize the associated variants, we used flow cytometry to measure TREM1 expression on monocytes.

Results: We provide evidence that an intronic variant, rs6910730(G) , in TREM1, is associated with an increased burden of neuritic plaques (p = 3.7 × 10(-4) ), diffuse plaques (p = 4.1 × 10(-3) ), and Aβ density (p = 2.6 × 10(-3) ) as well as an increased rate of cognitive decline (p = 5.3 × 10(-3) ). A variant upstream of TREM2, rs7759295(C) , is independently associated with an increased tau tangle density (p = 4.9 × 10(-4) ), an increased burden of neurofibrillary tangles (p = 9.1 × 10(-3) ), and an increased rate of cognitive decline (p = 2.3 × 10(-3) ). Finally, a cytometric analysis shows that the TREM1 rs6910730(G) allele is associated with decreased TREM1 expression on the surface of myeloid cells (p = 1.7 × 10(-3) ).

Interpretation: We provide evidence that 2 common variants within the TREM locus are associated with pathological features of AD and aging-related cognitive decline. Our evidence suggests that these variants are likely to be independent of known AD variants and that they may work through an alteration of myeloid cell function.
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http://dx.doi.org/10.1002/ana.24337DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4461024PMC
March 2015

Epigenomics in translational research.

Transl Res 2015 Jan 5;165(1):7-11. Epub 2014 Oct 5.

Department of Neurology, Brigham and Women's Hospital, Boston, Mass. Electronic address:

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http://dx.doi.org/10.1016/j.trsl.2014.09.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4533922PMC
January 2015

Polarization of the effects of autoimmune and neurodegenerative risk alleles in leukocytes.

Science 2014 May;344(6183):519-23

Program in Translational NeuroPsychiatric Genomics, Institute for the Neurosciences, Departments of Neurology and Psychiatry, Brigham and Women's Hospital, Boston, MA 02115, USA.

To extend our understanding of the genetic basis of human immune function and dysfunction, we performed an expression quantitative trait locus (eQTL) study of purified CD4(+) T cells and monocytes, representing adaptive and innate immunity, in a multi-ethnic cohort of 461 healthy individuals. Context-specific cis- and trans-eQTLs were identified, and cross-population mapping allowed, in some cases, putative functional assignment of candidate causal regulatory variants for disease-associated loci. We note an over-representation of T cell-specific eQTLs among susceptibility alleles for autoimmune diseases and of monocyte-specific eQTLs among Alzheimer's and Parkinson's disease variants. This polarization implicates specific immune cell types in these diseases and points to the need to identify the cell-autonomous effects of disease susceptibility variants.
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http://dx.doi.org/10.1126/science.1249547DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4910825PMC
May 2014

Interindividual variation in human T regulatory cells.

Proc Natl Acad Sci U S A 2014 Mar 7;111(12):E1111-20. Epub 2014 Mar 7.

Division of Immunology, Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115.

FOXP3(+) regulatory T (Treg) cells enforce immune self-tolerance and homeostasis, and variation in some aspects of Treg function may contribute to human autoimmune diseases. Here, we analyzed population-level Treg variability by performing genome-wide expression profiling of CD4(+) Treg and conventional CD4(+) T (Tconv) cells from 168 donors, healthy or with established type-1 diabetes (T1D) or type-2 diabetes (T2D), in relation to genetic and immunologic screening. There was a range of variability in Treg signature transcripts, some almost invariant, others more variable, with more extensive variability for genes that control effector function (ENTPD1, FCRL1) than for lineage-specification factors like FOXP3 or IKZF2. Network analysis of Treg signature genes identified coregulated clusters that respond similarly to genetic and environmental variation in Treg and Tconv cells, denoting qualitative differences in otherwise shared regulatory circuits whereas other clusters are coregulated in Treg, but not Tconv, cells, suggesting Treg-specific regulation of genes like CTLA4 or DUSP4. Dense genotyping identified 110 local genetic variants (cis-expression quantitative trait loci), some of which are specifically active in Treg, but not Tconv, cells. The Treg signature became sharper with age and with increasing body-mass index, suggesting a tuning of Treg function with repertoire selection and/or chronic inflammation. Some Treg signature transcripts correlated with FOXP3 mRNA and/or protein, suggesting transcriptional or posttranslational regulatory relationships. Although no single transcript showed significant association to diabetes, overall expression of the Treg signature was subtly perturbed in T1D, but not T2D, patients.
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http://dx.doi.org/10.1073/pnas.1401343111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970507PMC
March 2014

CD33: increased inclusion of exon 2 implicates the Ig V-set domain in Alzheimer's disease susceptibility.

Hum Mol Genet 2014 May 30;23(10):2729-36. Epub 2013 Dec 30.

Program in Translational NeuroPsychiatric Genomics, Institute for the Neurosciences, Departments of Neurology and Psychiatry, Brigham and Women's Hospital, Boston, MA 02115, USA.

We previously demonstrated that the Alzheimer's disease (AD) associated risk allele, rs3865444(C), results in a higher surface density of CD33 on monocytes. Here, we find alternative splicing of exon 2 to be the primary mechanism of the genetically driven differential expression of CD33 protein. We report that the risk allele, rs3865444(C), is associated with greater cell surface expression of CD33 in both subjects of European and African-American ancestry and that there is a single haplotype influencing CD33 surface expression. A meta-analysis of the two populations narrowed the number of significant SNPs in high linkage disequilibrium (LD) (r(2) > 0.8) with rs3865444 to just five putative causal variants associated with increased protein expression. Using gene expression data from flow-sorted CD14(+)CD16(-) monocytes from 398 healthy subjects of three populations, we show that the rs3865444(C) risk allele is strongly associated with greater expression of CD33 exon 2 (pMETA = 2.36 × 10(-60)). Western blotting confirms increased protein expression of the full-length CD33 isoform containing exon 2 relative to the rs3865444(C) allele (P < 0.0001). Of the variants in strong LD with rs3865444, rs12459419, which is located in a putative SRSF2 splice site of exon 2, is the most likely candidate to mediate the altered alternative splicing of CD33's Immunoglobulin V-set domain 2 and ultimately influence AD susceptibility.
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http://dx.doi.org/10.1093/hmg/ddt666DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3990171PMC
May 2014

Common risk alleles for inflammatory diseases are targets of recent positive selection.

Am J Hum Genet 2013 Apr 21;92(4):517-29. Epub 2013 Mar 21.

Program in Translational NeuroPsychiatric Genomics, Department of Neurology, Brigham & Women's Hospital, Boston, MA 02115, USA.

Genome-wide association studies (GWASs) have identified hundreds of loci harboring genetic variation influencing inflammatory-disease susceptibility in humans. It has been hypothesized that present day inflammatory diseases may have arisen, in part, due to pleiotropic effects of host resistance to pathogens over the course of human history, with significant selective pressures acting to increase host resistance to pathogens. The extent to which genetic factors underlying inflammatory-disease susceptibility has been influenced by selective processes can now be quantified more comprehensively than previously possible. To understand the evolutionary forces that have shaped inflammatory-disease susceptibility and to elucidate functional pathways affected by selection, we performed a systems-based analysis to integrate (1) published GWASs for inflammatory diseases, (2) a genome-wide scan for signatures of positive selection in a population of European ancestry, (3) functional genomics data comprised of protein-protein interaction networks, and (4) a genome-wide expression quantitative trait locus (eQTL) mapping study in peripheral blood mononuclear cells (PBMCs). We demonstrate that loci for inflammatory-disease susceptibility are enriched for genomic signatures of recent positive natural selection, with selected loci forming a highly interconnected protein-protein interaction network. Further, we identify 21 loci for inflammatory-disease susceptibility that display signatures of recent positive selection, of which 13 also show evidence of cis-regulatory effects on genes within the associated locus. Thus, our integrated analyses highlight a set of susceptibility loci that might subserve a shared molecular function and has experienced selective pressure over the course of human history; today, these loci play a key role in influencing susceptibility to multiple different inflammatory diseases, in part through alterations of gene expression in immune cells.
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http://dx.doi.org/10.1016/j.ajhg.2013.03.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3617371PMC
April 2013