@article{small_akhtari_green_havener_sikes_quintanhila_gonzalez_reif_motsinger-reif_mcleod_et al._2023, title={Pharmacogenomic Analyses Implicate B Cell Developmental Status and MKL1 as Determinants of Sensitivity toward Anti-CD20 Monoclonal Antibody Therapy}, volume={12}, ISSN={["2073-4409"]}, url={https://doi.org/10.3390/cells12121574}, DOI={10.3390/cells12121574}, abstractNote={Monoclonal antibody (mAb) therapy directed against CD20 is an important tool in the treatment of B cell disorders. However, variable patient response and acquired resistance remain important clinical challenges. To identify genetic factors that may influence sensitivity to treatment, the cytotoxic activity of three CD20 mAbs: rituximab; ofatumumab; and obinutuzumab, were screened in high-throughput assays using 680 ethnically diverse lymphoblastoid cell lines (LCLs) followed by a pharmacogenomic assessment. GWAS analysis identified several novel gene candidates. The most significant SNP, rs58600101, in the gene MKL1 displayed ethnic stratification, with the variant being significantly more prevalent in the African cohort and resulting in reduced transcript levels as measured by qPCR. Functional validation of MKL1 by shRNA-mediated knockdown of MKL1 resulted in a more resistant phenotype. Gene expression analysis identified the developmentally associated TGFB1I1 as the most significant gene associated with sensitivity. qPCR among a panel of sensitive and resistant LCLs revealed immunoglobulin class-switching as well as differences in the expression of B cell activation markers. Flow cytometry showed heterogeneity within some cell lines relative to surface Ig isotype with a shift to more IgG+ cells among the resistant lines. Pretreatment with prednisolone could partly reverse the resistant phenotype. Results suggest that the efficacy of anti-CD20 mAb therapy may be influenced by B cell developmental status as well as polymorphism in the MKL1 gene. A clinical benefit may be achieved by pretreatment with corticosteroids such as prednisolone followed by mAb therapy.}, number={12}, journal={CELLS}, author={Small, George W. and Akhtari, Farida S. and Green, Adrian J. and Havener, Tammy M. and Sikes, Michael and Quintanhila, Julia and Gonzalez, Ricardo D. and Reif, David M. and Motsinger-Reif, Alison A. and McLeod, Howard L. and et al.}, year={2023}, month={Jun} }
 @article{harris_sikes_bergman_goller_hasley_sjogren_ramirez_gordy_2022, title={Hands-on immunology: Engaging learners of all ages through tactile teaching tools}, volume={13}, ISSN={["1664-302X"]}, url={http://dx.doi.org/10.3389/fmicb.2022.966282}, DOI={10.3389/fmicb.2022.966282}, abstractNote={Ensuring the public has a fundamental understanding of human–microbe interactions, immune responses, and vaccines is a critical challenge in the midst of a pandemic. These topics are commonly taught in undergraduate- and graduate-level microbiology and immunology courses; however, creating engaging methods of teaching these complex concepts to students of all ages is necessary to keep younger students interested when science seems hard. Building on the Tactile Teaching Tools with Guided Inquiry Learning (TTT-GIL) method we used to create an interactive lac operon molecular puzzle, we report here two TTT-GIL activities designed to engage diverse learners from middle schoolers to masters students in exploring molecular interactions within the immune system. By pairing physical models with structured activities built on the constructivist framework of Process-Oriented Guided Inquiry Learning (POGIL), TTT-GIL activities guide learners through their interaction with the model, using the Learning Cycle to facilitate construction of new concepts. Moreover, TTT-GIL activities are designed utilizing Universal Design for Learning (UDL) principles to include all learners through multiple means of engagement, representation, and action. The TTT-GIL activities reported here include a web-enhanced activity designed to teach concepts related to antibody–epitope binding and specificity to deaf and hard-of-hearing middle and high school students in a remote setting and a team-based activity that simulates the evolution of the Major Histocompatibility Complex (MHC) haplotype of a population exposed to pathogens. These activities incorporate TTT-GIL to engage learners in the exploration of fundamental immunology concepts and can be adapted for use with learners of different levels and educational backgrounds.}, journal={FRONTIERS IN MICROBIOLOGY}, publisher={Frontiers Media SA}, author={Harris, Felix R. and Sikes, Michael L. and Bergman, Michael and Goller, Carlos C. and Hasley, Andrew O. and Sjogren, Caroline A. and Ramirez, Melissa V. and Gordy, Claire L.}, year={2022}, month={Aug} }
 @misc{azcarate-peril_sikes_bruno-barcena_2011, title={The intestinal microbiota, gastrointestinal environment and colorectal cancer: a putative role for probiotics in prevention of colorectal cancer?}, volume={301}, ISSN={["1522-1547"]}, url={http://europepmc.org/abstract/med/21700901}, DOI={10.1152/ajpgi.00110.2011}, abstractNote={Colorectal cancer (CRC) is the third most commonly diagnosed cancer in the United States, and, even though 5–15% of the total CRC cases can be attributed to individual genetic predisposition, environmental factors could be considered major factors in susceptibility to CRC. Lifestyle factors increasing the risks of CRC include elevated body mass index, obesity, and reduced physical activity. Additionally, a number of dietary elements have been associated with higher or lower incidence of CRC. In this context, it has been suggested that diets high in fruit and low in meat might have a protective effect, reducing the incidence of colorectal adenomas by modulating the composition of the normal nonpathogenic commensal microbiota. In addition, it has been demonstrated that changes in abundance of taxonomic groups have a profound impact on the gastrointestinal physiology, and an increasing number of studies are proposing that the microbiota mediates the generation of dietary factors triggering colon cancer. High-throughput sequencing and molecular taxonomic technologies are rapidly filling the knowledge gaps left by conventional microbiology techniques to obtain a comprehensive catalog of the human intestinal microbiota and their associated metabolic repertoire. The information provided by these studies will be essential to identify agents capable of modulating the massive amount of gut bacteria in safe noninvasive manners to prevent CRC. Probiotics, defined as “live microorganisms which, when administered in adequate amounts, confer a health benefit on the host” ( 219 ), are capable of transient modulation of the microbiota, and their beneficial effects include reinforcement of the natural defense mechanisms and protection against gastrointestinal disorders. Probiotics have been successfully used to manage infant diarrhea, food allergies, and inflammatory bowel disease; hence, the purpose of this review was to examine probiotic metabolic activities that may have an effect on the prevention of CRC by scavenging toxic compounds or preventing their generation in situ. Additionally, a brief consideration is given to safety evaluation and production methods in the context of probiotics efficacy.}, number={3}, journal={AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY}, author={Azcarate-Peril, M. Andrea and Sikes, Michael and Bruno-Barcena, Jose M.}, year={2011}, month={Sep}, pages={G401–G424} }
 @article{troxell_sikes_fink_vazquez-torres_jones-carson_hassan_2011, title={Fur Negatively Regulates hns and Is Required for the Expression of HilA and Virulence in Salmonella enterica Serovar Typhimurium}, volume={193}, ISSN={["1098-5530"]}, DOI={10.1128/jb.00942-10}, abstractNote={ABSTRACT
          
            Iron is an essential element for the survival of living cells. However, excess iron is toxic, and its uptake is exquisitely regulated by the ferric uptake regulator, Fur. In
            Salmonella
            , the
            Salmonella
            pathogenicity island 1 (SPI-1) encodes a type three secretion system, which is required for invasion of host epithelial cells in the small intestine. A major activator of SPI-1 is HilA, which is encoded within SPI-1. One known regulator of
            hilA
            is Fur. The mechanism of
            hilA
            regulation by Fur is unknown. We report here that Fur is required for virulence in
            Salmonella enterica
            serovar Typhimurium and that Fur is required for the activation of
            hilA
            , as well as of other HilA-dependent genes,
            invF
            and
            sipC
            . The Fur-dependent regulation of
            hilA
            was independent of PhoP, a known repressor of
            hilA
            . Instead, the expression of the gene coding for the histone-like protein,
            hns
            , was significantly derepressed in the
            fur
            mutant. Indeed, the activation of
            hilA
            by Fur was dependent on 28 nucleotides located upstream of
            hns
            . Moreover, we used chromatin immunoprecipitation to show that Fur bound,
            in vivo
            , to the upstream region of
            hns
            in a metal-dependent fashion. Finally, deletion of
            fur
            in an
            hns
            mutant resulted in Fur-independent activation of
            hilA
            . In conclusion, Fur activates
            hilA
            by repressing the expression of
            hns
            .
          }, number={2}, journal={JOURNAL OF BACTERIOLOGY}, author={Troxell, Bryan and Sikes, Michael L. and Fink, Ryan C. and Vazquez-Torres, Andres and Jones-Carson, Jessica and Hassan, Hosni M.}, year={2011}, month={Jan}, pages={497–505} }
 @article{oyegunwa_sikes_wilson_scholle_laster_2010, title={Tetra-O-methyl nordihydroguaiaretic acid (Terameprocol) inhibits the NF-kappa B-dependent transcription of TNF-alpha and MCP-1/CCL2 genes by preventing RelA from binding its cognate sites on DNA}, volume={7}, ISSN={["1476-9255"]}, DOI={10.1186/1476-9255-7-59}, abstractNote={Abstract
          
            Background
            Tetra-O-methyl nordihydroguaiaretic acid, also known as terameprocol (TMP), is a naturally occurring phenolic compound found in the resin of the creosote bush. We have shown previously that TMP will suppress production of certain inflammatory cytokines, chemokines and lipids from macrophages following stimulation with LPS or infection with H1N1 influenza virus. In this study our goal was to elucidate the mechanism underlying TMP-mediated suppression of cytokine and chemokine production. We focused our investigations on the response to LPS and the NF-κB protein RelA, a transcription factor whose activity is critical to LPS-responsiveness.
          
          
            Methods
            Reporter assays were performed with HEK293 cells overexpressing either TLR-3, -4, or -8 and a plasmid containing the luciferase gene under control of an NF-κB response element. Cells were then treated with LPS, poly(I:C), or resiquimod, and/or TMP, and lysates measured for luciferase activity.
            RAW 264.7 cells treated with LPS and/or TMP were used in ChIP and EMSA assays. For ChIP assays, chromatin was prepared and complexes precipitated with anti-NF-κB RelA Ab. Cross-links were reversed, DNA purified, and sequence abundance determined by Q-PCR. For EMSA assays, nuclear extracts were incubated with radiolabeled probes, analyzed by non-denaturing PAGE and visualized by autoradiography.
            RAW 264.7 cells treated with LPS and/or TMP were also used in fluorescence microscopy and western blot experiments. Translocation experiments were performed using a primary Ab to NF-κB RelA and a fluorescein-conjugated secondary Ab. Western blots were performed using Abs to IκB-α and phospho-IκB-α. Bands were visualized by chemiluminescence.
          
          
            Results
            In reporter assays with TLR-3, -4, and -8 over-expressing cells, TMP caused strong inhibition of NF-κB-dependent transcription.
            ChIP assays showed TMP caused virtually complete inhibition of RelA binding in vivo to promoters for the genes for TNF-α, MCP-1/CCL2, and RANTES/CCL5 although the LPS-dependent synthesis of IκB-α was not inhibited. EMSA assays did not reveal an effect of TMP on the binding of RelA to naked DNA templates in vitro.
            TMP did not inhibit the nuclear translocation of NF-κB RelA nor the phosphorylation of IκB-α.
          
          
            Conclusion
            TMP acts indirectly as an inhibitor of NF-κB-dependent transcription by preventing RelA from binding the promoters of certain key cytokine and chemokine genes.
          }, journal={JOURNAL OF INFLAMMATION-LONDON}, author={Oyegunwa, Akinbolade O. and Sikes, Michael L. and Wilson, Jason R. and Scholle, Frank and Laster, Scott M.}, year={2010}, month={Dec} }
 @misc{sikes_mcmillan_bradshaw_2010, title={The Center of Accessibility: D beta Control of V(D)J Recombination}, volume={58}, ISSN={["0004-069X"]}, DOI={10.1007/s00005-010-0101-2}, abstractNote={Developmental patterning of antigen receptor gene assembly in lymphocyte precursors correlates with decondensation of the chromatin surrounding individual gene segments. Ongoing V(D)J recombination is associated with hyperacetylation of histones H3 and H4 and the expression of sterile germline transcripts across the region of recombinational accessibility. Likewise, histone acetyltransferase and SWI/SNF chromatin remodeling complexes each appear to be required for recombination, and the PHD-finger of RAG-2 preferentially associates with recombination signal sequence (RSS) chromatin that contains H3 trimethylated on lysine 4. However, the regulatory mechanisms that direct chromatin alteration and rearrangement have proven elusive, due in large part to the interdependency of individual stages in gene activation, our limited understanding of functional significance of changes to the histone code, and the difficulty of modeling recombinational accessibility in existing experimental systems. Examining Tcrb assembly in developing thymocytes, we review the central roles of RSS elements and germline promoters as foci for epigenetic reorganization of recombinationally accessible gene segments in light of recent findings and persistent questions.}, number={6}, journal={ARCHIVUM IMMUNOLOGIAE ET THERAPIAE EXPERIMENTALIS}, author={Sikes, Michael L. and McMillan, Ruth E. and Bradshaw, Justin M.}, year={2010}, month={Dec}, pages={427–433} }
 @article{sikes_bradshaw_ivory_lunsford_mcmillan_morrison_2009, title={A streamlined method for rapid and sensitive chromatin immunoprecipitation}, volume={344}, ISSN={["0022-1759"]}, DOI={10.1016/j.jim.2009.03.007}, abstractNote={We report a streamlined procedure to efficiently carry samples from chromatin to qPCR-compatible DNA in as little as 4 h. We use this streamlined ChIP to quantify histone H3 modifications at active (cad) and repressed (T early alpha) promoters in a Rag1-deficient pro-T cell line after 1–2 h IP. We further show that the protocol readily quantified histone modifications in chromatin from 104 Rag-deficient DN thymocytes. Taken together, these data outline a simple, cost-effective procedure for efficient ChIP analysis.}, number={1}, journal={JOURNAL OF IMMUNOLOGICAL METHODS}, author={Sikes, Michael L. and Bradshaw, Justin M. and Ivory, Wendell T. and Lunsford, Jessica L. and McMillan, Ruth E. and Morrison, Clayton R.}, year={2009}, month={May}, pages={58–63} }
 @article{taveirne_sikes_olson_2009, title={Molybdenum and tungsten in Campylobacter jejuni: their physiological role and identification of separate transporters regulated by a single ModE-like protein}, volume={74}, ISSN={["1365-2958"]}, DOI={10.1111/j.1365-2958.2009.06901.x}, abstractNote={Summary Campylobacter jejuni is an important human pathogen that causes millions of cases of food‐borne enteritis each year. The C. jejuni respiratory chain is highly branched and contains at least four enzymes predicted to contain a metal binding pterin (MPT), with the metal being either molybdenum or tungsten. Also predicted are two separate transport systems, one for molybdenum encoded by modABC and a second for tungsten encoded by tupABC. Both transport systems were mutated and the activities of the four predicted MPT‐containing enzymes were assayed in the presence of molybdenum and tungsten in wild‐type and mod and tup backgrounds. Results indicate that mod is primarily a molybdenum transporter that can also transport tungsten, while tup is a tungsten‐specific transporter. The MPT‐containing enzymes nitrate reductase, sulphite oxidase, and SN oxide reductase are strict molybdoenzymes while formate dehydrogenase prefers tungsten. A ModE‐like protein regulates both transporters, repressing mod in the presence of both molybdenum and tungsten and tup only in the presence of tungsten. Like other ModE proteins, the C. jejuni ModE binds DNA through a helix–turn–helix DNA binding domain, but unlike other members of the ModE family it does not have a metal binding domain.}, number={3}, journal={MOLECULAR MICROBIOLOGY}, author={Taveirne, Michael E. and Sikes, Michael L. and Olson, Jonathan W.}, year={2009}, month={Nov}, pages={758–771} }
 @article{mcmillan_sikes_2009, title={Promoter activity 5 ' of D beta 2 is coordinated by E47, Runx1, and GATA-3}, volume={46}, ISSN={["0161-5890"]}, DOI={10.1016/j.molimm.2009.06.013}, abstractNote={V(D)J recombination involves the stepwise assembly of B and T cell receptor genes as lymphocytes progress through the early stages of development. While the mechanisms that restrict each step in recombination to its appropriate developmental stage are largely unknown, they share many of the components that regulate transcription. For example, enhancer-dependent modifications in histone acetylation and methylation are essential for both germline transcription and rearrangement of antigen receptor genes. Promoters positioned proximal to individual D and J gene segments in Tcra, Tcrb, Tcrd, IgH, and Igk also contribute to antigen receptor gene assembly, though their effects appear more localized than those of enhancers. Tcrb assembly initiates with D-to-J joining at each of the two D–J–C gene segment clusters in DN1/2 thymocytes. DJ joints are fused with Vβ elements to complete Tcrb recombination in DN3 cells. We have previously shown that Dβ2 is flanked by upstream and downstream promoters, with the 5′ promoter being held inactive until D-to-J recombination deletes the NFκB-dependent 3′ promoter. We now report that activity of the 5′ promoter reflects a complex interplay among Runx1, GATA-3, and E47 transcription factors. In particular, while multiple E47 and Runx1 binding sites clustered near the Dβ2 5′RS and overlapping inr elements define the core 5′PDβ2, they act in concert with an array of upstream GATA-3 sites to overcome the inhibitory effects of a 110 bp distal polypurine·polypyrimidine (R·Y) tract. The dependence of 5′PDβ2 on E47 is consistent with the reported role of E proteins in post-DN1 thymocyte development and V-to-DJβ recombination.}, number={15}, journal={MOLECULAR IMMUNOLOGY}, author={McMillan, Ruth E. and Sikes, Michael L.}, year={2009}, month={Sep}, pages={3009–3017} }
 @article{mcmillan_sikes_2008, title={Differential activation of dual promoters alters D beta 2 germline transcription during thymocyte development}, volume={180}, ISSN={["0022-1767"]}, DOI={10.4049/jimmunol.180.5.3218}, abstractNote={Abstract
               Ag receptor genes are assembled through somatic rearrangements of V, D, and J gene segments. This process is directed in part by transcriptional enhancers and promoters positioned within each gene locus. Whereas enhancers coordinate reorganization of large chromatin stretches, promoters are predicted to facilitate the accessibility of proximal downstream gene segments. In TCR β locus, rearrangement initiates at two D-J cassettes, each of which exhibits transcriptional activity coincident with DJ rearrangement in CD4/CD8 double-negative pro-T cells. Consistent with a model of promoter-facilitated recombination, assembly of the DJβ1 cassette is dependent on a Dβ1 promoter (PDβ1) positioned immediately 5′ of the D. Assembly of DJβ2 proceeds independent from that of DJβ1, albeit with less efficiency. To gain insight into the mechanisms that selectively alter D usage, we have defined transcriptional regulation at Dβ2. We find that both DJβ cassettes generate germline messages in murine CD44+CD25− double-negative 1 cells. However, transcription of unrearranged DJβ2 initiates at multiple sites 400–550 bp downstream of the Dβ2. Unexpectedly, loci from which germline promoter activity has been deleted by DJ rearrangement redirect transcription to sites immediately 5′ of the new DJβ2 joint. Our analyses suggest that 3′-PDβ2 activity is largely controlled by NF-κB RelA, whereas 5′-PDβ2 activity directs germline transcription of DJβ2 joints from initiator elements 76 bp upstream of the Dβ2 5′ recombination signal sequence. The unique organization and timing of Dβ2 promoter activity are consistent with a model in which promoter placement selectively regulates the rearrangement potential of Dβ2 during TCR β locus assembly.}, number={5}, journal={JOURNAL OF IMMUNOLOGY}, author={McMillan, Ruth E. and Sikes, Michael L.}, year={2008}, month={Mar}, pages={3218–3228} }