Publications by authors named "Vipul Shukla"

15 Publications

  • Page 1 of 1

HMCES Functions in the Alternative End-Joining Pathway of the DNA DSB Repair during Class Switch Recombination in B Cells.

Mol Cell 2020 01 2;77(2):384-394.e4. Epub 2019 Dec 2.

National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD 20894, USA. Electronic address:

HMCES (5hmC binding, embryonic stem cell-specific-protein), originally identified as a protein capable of binding 5-hydroxymethylcytosine (5hmC), an epigenetic modification generated by TET proteins, was previously reported to covalently crosslink to DNA at abasic sites via a conserved cysteine. We show here that Hmces-deficient mice display normal hematopoiesis without global alterations in 5hmC. HMCES specifically enables DNA double-strand break repair through the microhomology-mediated alternative-end-joining (Alt-EJ) pathway during class switch recombination (CSR) in B cells, and HMCES deficiency leads to a significant defect in CSR. HMCES mediates Alt-EJ through its SOS-response-associated-peptidase domain (SRAPd), a function that requires DNA binding but is independent of its autopeptidase and DNA-crosslinking activities. We show that HMCES is recruited to switch regions of the immunoglobulin locus and provide a potential structural basis for the interaction of HMCES with long DNA overhangs generated by Alt-EJ during CSR. Our studies provide further evidence for a specialized role for HMCES in DNA repair.
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http://dx.doi.org/10.1016/j.molcel.2019.10.031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6980713PMC
January 2020

TET enzymes augment activation-induced deaminase (AID) expression via 5-hydroxymethylcytosine modifications at the superenhancer.

Sci Immunol 2019 04;4(34)

Division of Signaling and Gene Expression, La Jolla Institute, San Diego, CA, USA.

TET enzymes are dioxygenases that promote DNA demethylation by oxidizing the methyl group of 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Here, we report a close correspondence between 5hmC-marked regions, chromatin accessibility and enhancer activity in B cells, and a strong enrichment for consensus binding motifs for basic region-leucine zipper (bZIP) transcription factors at TET-responsive genomic regions. Functionally, Tet2 and Tet3 regulate class switch recombination (CSR) in murine B cells by enhancing expression of , which encodes the activation-induced cytidine deaminase (AID) enzyme essential for CSR. TET enzymes deposit 5hmC, facilitate DNA demethylation, and maintain chromatin accessibility at two TET-responsive enhancer elements, and , located within a superenhancer in the locus. Our data identify the bZIP transcription factor, ATF-like (BATF) as a key transcription factor involved in TET-dependent expression. 5hmC is not deposited at in activated -deficient B cells, indicating that BATF facilitates TET recruitment to this enhancer. Our study emphasizes the importance of TET enzymes for bolstering AID expression and highlights 5hmC as an epigenetic mark that captures enhancer dynamics during cell activation.
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http://dx.doi.org/10.1126/sciimmunol.aau7523DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6599614PMC
April 2019

Regulation of MAPK signaling and implications in chronic lymphocytic leukemia.

Leuk Lymphoma 2018 07 7;59(7):1565-1573. Epub 2017 Sep 7.

c Department of Genetics Cell Biology and Anatomy , University of Nebraska Medical Centre , Omaha , NE , USA.

Chronic lymphocytic leukemia (CLL) is a heterogeneous B cell malignancy that still remains incurable. Recent studies have highlighted cellular and non-cellular components of the tissue microenvironment in CLL that help nurture the growth of leukemic cells by providing the necessary stimuli for their proliferation and survival. The diverse stimuli in the specialized tissue microenvironment of CLL lead to constitutive activation of several signaling pathways that includes B cell receptor signaling and the associated mitogen-activated protein kinase (MAPK) signaling. Recent findings have described aberrant activation of MAPK signaling and its interactions with other cellular signaling pathways in the pathogenesis of CLL. These studies have shed light on the deregulated molecular mechanisms contributing to hyperactivation of MAPK signaling and provided avenues for therapeutic options for aggressive CLL. In this review, we describe and discuss the current status of our understanding into the role of MAPK signaling in the pathogenesis of CLL.
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http://dx.doi.org/10.1080/10428194.2017.1370548DOI Listing
July 2018

Addressing the healthcare needs of older Lesbian, Gay, Bisexual, and Transgender patients in medical school curricula: a call to action.

Med Educ Online 2017 ;22(1):1320933

c Department of Family Medicine, College of Medicine and Life Sciences , University of Toledo , Toledo , OH , USA.

Medical students matriculating in the coming years will be faced with treating an expansive increase in the population of older lesbian, gay, bisexual, and transgender (LGBT) patients. While these patients face healthcare concerns similar to their non-LGBT aging peers, the older LGBT community has distinct healthcare needs and faces well-documented healthcare disparities. In order to reduce these healthcare barriers, medical school curricula must prepare and educate future physicians to treat this population while providing high quality, culturally-competent care. This article addresses some of the unique healthcare needs of the aging LGBT population with an emphasis on social concerns and healthcare disparities. It provides additional curricular recommendations to aid in the progressive augmentation of medical school curricula.

Abbreviations: Liaison Committee on Medical Education (LCME); LGBT: Lesbian, gay, bisexual, transgender.
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http://dx.doi.org/10.1080/10872981.2017.1320933DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5419296PMC
March 2018

Violent Video Games Exposed: A Blow by Blow Account of Senseless Violence in Games.

J Psychol 2017 Jan 23;151(1):76-87. Epub 2016 Sep 23.

a University of Toledo.

Violent video game (VVG) use has repeatedly been found to be associated with hostile expectations about others, desensitization to violence, decreased empathy and prosocial behavior, and aggressive thoughts and behaviors. Although these research findings have been widely publicized, VVGs remain the most extensively played games and represent a multi-billion dollar industry. Although VVGs are typically rated "mature," indicating they are not suitable for youths, they are often purchased for youths. This may be in part because there is currently no system available to consumers that thoroughly describes the content of video games, and much of the public is unaware of the types of violence that characterize game play. The purpose of this paper is to describe the violent content of some of the top VVGs, based on sales. For the purposes of this issue, acts of senseless, unprovoked violence will be described in detail.
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http://dx.doi.org/10.1080/00223980.2016.1226744DOI Listing
January 2017

Interferon regulatory factor 4 attenuates Notch signaling to suppress the development of chronic lymphocytic leukemia.

Oncotarget 2016 Jul;7(27):41081-41094

Department of Genetics Cell Biology and Anatomy, University of Nebraska Medical Center, Omaha, NE, USA.

Molecular pathogenesis of Chronic Lymphocytic Leukemia (CLL) is not fully elucidated. Genome wide association studies have linked Interferon Regulatory Factor 4 (IRF4) to the development of CLL. We recently established a causal relationship between low levels of IRF4 and development of CLL. However, the molecular mechanism through which IRF4 suppresses CLL development remains unclear. Deregulation of Notch signaling pathway has been identified as one of the most recurrent molecular anomalies in the pathogenesis of CLL. Yet, the role of Notch signaling as well as its regulation during CLL development remains poorly understood. Previously, we demonstrated that IRF4 deficient mice expressing immunoglobulin heavy chain Vh11 (IRF4-/-Vh11) developed spontaneous CLL with complete penetrance. In this study, we show that elevated Notch2 expression and the resulting hyperactivation of Notch signaling are common features of IRF4-/-Vh11 CLL cells. Our studies further reveal that Notch signaling is indispensable for CLL development in the IRF4-/-Vh11 mice. Moreover, we identify E3 ubiquitin ligase Nedd4, which targets Notch for degradation, as a direct target of IRF4 in CLL cells and their precursors. Collectively, our studies provide the first in vivo evidence for an essential role of Notch signaling in the development of CLL and establish IRF4 as a critical regulator of Notch signaling during CLL development.
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http://dx.doi.org/10.18632/oncotarget.9596DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5173044PMC
July 2016

Promoter Hypomethylation and Expression Is Conserved in Mouse Chronic Lymphocytic Leukemia Induced by Decreased or Inactivated Dnmt3a.

Cell Rep 2016 05 28;15(6):1190-201. Epub 2016 Apr 28.

Department of Genetics, Cell Biology, and Anatomy, University of Nebraska Medical Center, Omaha, NE 68198, USA; Eppley Institute for Research in Cancer and Allied Diseases, Fred and Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE 68198, USA; Center for Leukemia and Lymphoma Research, University of Nebraska Medical Center, Omaha, NE 68198, USA. Electronic address:

DNA methyltransferase 3a (DNMT3A) catalyzes the formation of 5-methyl-cytosine in mammalian genomic DNA, and it is frequently mutated in human hematologic malignancies. Bi-allelic loss of Dnmt3a in mice results in leukemia and lymphoma, including chronic lymphocytic leukemia (CLL). Here, we investigate whether mono-allelic loss of Dnmt3a is sufficient to induce disease. We show that, by 16 months of age, 65% of Dnmt3a(+/-) mice develop a CLL-like disease, and 15% of mice develop non-malignant myeloproliferation. Genome-wide methylation analysis reveals that reduced Dnmt3a levels induce promoter hypomethylation at similar loci in Dnmt3a(+/-) and Dnmt3a(Δ/Δ) CLL, suggesting that promoters are particularly sensitive to Dnmt3a levels. Gene expression analysis identified 26 hypomethylated and overexpressed genes common to both Dnmt3a(+/-) and Dnmt3a(Δ/Δ) CLL as putative oncogenic drivers. Our data provide evidence that Dnmt3a is a haplo-insufficient tumor suppressor in CLL and highlights the importance of deregulated molecular events in disease pathogenesis.
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http://dx.doi.org/10.1016/j.celrep.2016.04.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4864108PMC
May 2016

Sprouty 2: a novel attenuator of B-cell receptor and MAPK-Erk signaling in CLL.

Blood 2016 05 25;127(19):2310-21. Epub 2016 Jan 25.

Department of Genetics, Cell Biology and Anatomy.

Clinical heterogeneity is a major barrier to effective treatment of chronic lymphocytic leukemia (CLL). Emerging evidence suggests that constitutive activation of various signaling pathways like mitogen-activated protein kinase-extracellular signal-regulated kinase (MAPK-Erk) signaling plays a role in the heterogeneous clinical outcome of CLL patients. In this study, we have investigated the role of Sprouty (SPRY)2 as a negative regulator of receptor and nonreceptor tyrosine kinase signaling in the pathogenesis of CLL. We show that SPRY2 expression is significantly decreased in CLL cells, particularly from poor-prognosis patients compared with those from good-prognosis patients. Overexpression of SPRY2 in CLL cells from poor-prognosis patients increased their apoptosis. Conversely, downregulation of SPRY2 in CLL cells from good-prognosis patients resulted in increased proliferation. Furthermore, CLL cells with low SPRY2 expression grew more rapidly in a xenograft model of CLL. Strikingly, B-cell-specific transgenic overexpression of spry2 in mice led to a decrease in the frequency of B1 cells, the precursors of CLL cells in rodents. Mechanistically, we show that SPRY2 attenuates the B-cell receptor (BCR) and MAPK-Erk signaling by binding to and antagonizing the activities of RAF1, BRAF, and spleen tyrosine kinase (SYK) in normal B cells and CLL cells. We also show that SPRY2 is targeted by microRNA-21, which in turn leads to increased activity of Syk and Erk in CLL cells. Taken together, these results establish SPRY2 as a critical negative regulator of BCR-mediated MAPK-Erk signaling in CLL, thereby providing one of the molecular mechanisms to explain the clinical heterogeneity of CLL.
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http://dx.doi.org/10.1182/blood-2015-09-669317DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4865588PMC
May 2016

Alternative antiretroviral therapy formulations for patients unable to swallow solid oral dosage forms.

Am J Health Syst Pharm 2015 Sep;72(18):1555-65

Joan M. Duggan, M.D., FACP, FIDSA, AAHIVS, is Infectious Diseases Specialist; and Barbara Akpanudo, M.S.N., M.P.H., CNP, is Family Nurse Practitioner, Division of Infectious Diseases, Department of Medicine, University of Toledo Medical Center (UTMC), Toledo, OH. Vipul Shukla, B.S., is a medical student; and Glen Gutterson, B.S., is a medical student, College of Medicine, University of Toledo. Lindsey Eitniear, Pharm.D., is Clinical Pharmacist, UTMC. Eric G. Sahloff, Pharm.D., AAHIVP, is Associate Professor of Pharmacy Practice, College of Pharmacy and Pharmaceutical Sciences, University of Toledo.

Purpose: Evidence-based guidance is presented to assist clinicians in selecting alternative formulations of antiretroviral (ARV) agents for patients with human immunodeficiency virus (HIV) infection who are unable to swallow tablets or capsules.

Summary: The inability to take medications in standard oral dosage forms can be associated with nonadherence or the use of alternative administration strategies such as capsule or tablet breaking, crushing, or chewing. Patients with HIV infection require long-term ARV therapy to maintain viral suppression; ARV agents are predominately available as tablets and capsules that may pose swallowing difficulties for some patients. Using a variety of sources (the primary literature, pharmaceutical package inserts, and requests for unpublished data from drug manufacturers), available evidence on the bioavailability of ARV medications after disruption of the capsule or tablet matrix was reviewed; information on alternative formulations of ARV agents was also assessed. With several ARV agents, disruption of the solid oral dosage form by crushing, chewing, or breaking tablets or opening capsules prior to ingestion has been shown to result in altered bioavailability or pharmacokinetics and thus the potential for incomplete virological suppression, increased adverse effects, and suboptimal health outcomes.

Conclusion: Of the 33 single-agent ARV medications and combination ARV products in five classes available at the time of review, approximately half exist as powders, liquids, injectables, or chewable or dissolvable tablets. If alternative ARV formulations or administration methods are used, close monitoring for achievement of virological and immunologic success and potential toxicities is recommended.
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http://dx.doi.org/10.2146/ajhp140575DOI Listing
September 2015

IRF4 and IRF8: Governing the virtues of B Lymphocytes.

Front Biol (Beijing) 2014 Aug;9(4):269-282

Department of Genetics, Cell Biology & Anatomy. University of Nebraska Medical Center, Omaha, NE 68198.

Interferon Regulatory Factor 4 (IRF4) and IRF8 are critical regulators of immune system development and function. In B lymphocytes, IRF4 and IRF8 have been shown to control important events during their development and maturation including pre-B cell differentiation, induction of B cell tolerance pathways, marginal zone B cell development, germinal center reaction and plasma cell differentiation. Mechanistically, IRF4 and IRF8 are found to function redundantly to control certain stages of B cell development, but in other stages, they function nonredundantly to play distinct roles in B cell biology. In line with their essential roles in B cell development, deregulated expressions of IRF4 and IRF8 have been associated to the pathogenesis of several B cell malignancies and diseases. Recent studies have elucidated diverse transcriptional networks regulated by IRF4 and IRF8 at distinct B cell developmental stages and related malignancies. In this review we will discuss the recent advances for the roles of IRF4 and IRF8 during B cell development and associated diseases.
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http://dx.doi.org/10.1007/s11515-014-1318-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4261187PMC
August 2014

Barriers to Healthcare in the Transgender Community: A Case Report.

LGBT Health 2014 Sep 21;1(3):229-32. Epub 2014 Apr 21.

College of Medicine, University of Toledo-Health Science Campus , Toledo, Ohio.

The transgender community has many unique health needs that are often inadequately addressed by the current medical system. Healthcare workers may lack adequate training to deal with these unique issues, compromising care. Healthcare workers must understand the major barriers to healthcare and heath disparities experienced by the transgender community in order to work to overcome them. This article presents a case illustrating some of the issues that may compromise healthcare for the transgender community as well as measures taken by the providers in response to this case to improve caring for this population in the future.
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http://dx.doi.org/10.1089/lgbt.2013.0054DOI Listing
September 2014

A role for IRF8 in B cell anergy.

J Immunol 2013 Dec 11;191(12):6222-30. Epub 2013 Nov 11.

Department of Genetics, Cell Biology, and Anatomy, University of Nebraska Medical Center, Omaha, NE 68198.

B cell central tolerance is a process through which self-reactive B cells are removed from the B cell repertoire. Self-reactive B cells are generally removed by receptor editing in the bone marrow and by anergy induction in the periphery. IRF8 is a critical transcriptional regulator of immune system development and function. A recent study showed that marginal zone B cell and B1 B cell populations are dramatically increased in IRF8-deficient mice, indicating that there are B cell-developmental defects in the absence of IRF8. In this article, we report that mice deficient for IRF8 produced anti-dsDNA Abs. Using a hen egg lysozyme double-transgenic model, we further demonstrate that B cell anergy was breached in IRF8-deficient mice. Although anergic B cells in the IRF8-proficient background were blocked at the transitional stage of development, anergic B cells in the IRF8-deficient background were able to mature further, which allowed them to regain responses to Ag stimulation. Interestingly, our results show that IRF8-deficient B cells were more sensitive to Ag stimulation and were resistant to Ag-induced cell death. Moreover, our results show that IRF8 was expressed at a high level in the anergic B cells, and an elevated level of IRF8 promoted apoptosis in the transitional B cells. Thus, our findings reveal a previously unrecognized function of IRF8 in B cell anergy induction.
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http://dx.doi.org/10.4049/jimmunol.1301169DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3864091PMC
December 2013

A role for IRF4 in the development of CLL.

Blood 2013 Oct 7;122(16):2848-55. Epub 2013 Aug 7.

Department of Genetics, Cell Biology, and Anatomy, University of Nebraska Medical Center, Omaha, NE; and.

Interferon regulatory factor 4 (IRF4) is a critical transcriptional regulator of B-cell development and function. A recent genome-wide single-nucleotide polymorphism (SNP) association study identified IRF4 as a major susceptibility gene in chronic lymphocytic leukemia (CLL). Although the SNPs located in the IRF4 gene were linked to a downregulation of IRF4 in CLL patients, whether a low level of IRF4 is critical for CLL development remains unclear. In rodents, CLL cells are derived from B1 cells whose population is dramatically expanded in immunoglobulin heavy chain Vh11 knock-in mice. We bred a Vh11 knock-in allele into IRF4-deficient mice (IRF4(-/-)Vh11). Here, we report that IRF4(-/-)Vh11 mice develop spontaneous early-onset CLL with 100% penetrance. Further analysis shows that IRF4(-/-)Vh11 CLL cells proliferate predominantly in spleen and express high levels of Mcl-1. IRF4(-/-)Vh11 CLL cells are resistant to apoptosis but reconstitution of IRF4 expression in the IRF4(-/-)Vh11 CLL cells inhibits their survival. Thus, our study demonstrates for the first time a causal relationship between low levels of IRF4 and the development of CLL. Moreover, our findings establish IRF4(-/-)Vh11 mice as a novel mouse model of CLL that not only is valuable for dissecting molecular pathogenesis of CLL but could also be used for therapeutic purposes.
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http://dx.doi.org/10.1182/blood-2013-03-492769DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3798999PMC
October 2013

Accelerated development of chronic lymphocytic leukemia in New Zealand Black mice expressing a low level of interferon regulatory factor 4.

J Biol Chem 2013 Sep 29;288(37):26430-40. Epub 2013 Jul 29.

From the Department of Genetics, Cell Biology, and Anatomy, University of Nebraska Medical Center, Omaha, Nebraska 68198.

A recent genome-wide SNP association study identified IRF4 as a major susceptibility gene for chronic lymphocytic leukemia (CLL). Moreover, the SNPs located in the 3' UTR of the IRF4 gene have been linked to a down-regulation of IRF4. However, whether a low level of IRF4 is critical for CLL development remains unclear. New Zealand Black (NZB) mice are a naturally occurring, late-onset mouse model of CLL. To examine the role of a reduced level of IRF4 in CLL development, we generated, through breeding, IRF4 heterozygous mutant mice in the NZB background (NZB IRF4(+/-)). Our results show that CLL development is accelerated dramatically in the NZB IRF4(+/-) mice. The average onset of CLL in NZB mice is 12 months, but CLL cells can be detected in NZB IRF4(+/-) mice at 3 months of age. By 5 months of age, 80% of NZB IRF4(+/-) mice developed CLL. CLL cells are derived from B1 cells in mice. Interestingly, NZB IRF4(+/-) B1 cells exhibit prolonged survival, accelerated self-renewal, and defects in differentiation. Although NZB IRF4(+/-) CLL cells are resistant to apoptosis, high levels of IRF4 inhibit their survival. High levels of IRF4 also reduce the survival of MEC-1 human CLL cells. Our analysis further reveals that high levels of IRF4 suppress Akt activity and can do so without the IRF4 DNA binding domain. Thus, our findings reveal a causal relationship between a low level of IRF4 and the development of CLL and establish IRF4 as a novel regulator in the pathogenesis of CLL.
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http://dx.doi.org/10.1074/jbc.M113.475913DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3772189PMC
September 2013