Associate Professor of Cell and Developmental BiologyLab Homepage: http://www.brarlab.org/
The mechanisms that link cellular differentiation programs and dynamic gene regulation in complex eukaryotic systems remain mysterious. Such programs drive diverse and central biological processes including organismal development, immune function, disease progression, and meiosis. Our lab is focused on the molecular basis for the cellular remodeling accompanying meiosis, the highly conserved process by which gametes are produced.
We study meiosis because it is itself a biologically interesting and important process, but also because it serves as a tractable model for the complex cellular changes that accompany many types of differentiation. We are interested in understanding the fundamental mechanisms by which a cell achieves such changes. Towards this end, we use high-throughput and classical genetic and molecular approaches to study the role of pervasive short protein synthesis in meiosis, specialized regulation of meiotic translation, and the role of several prominent and diverse stress response pathways in driving cells through the meiotic program.
We have globally probed the regulation of the comprehensive cellular restructuring underlying meiosis. Ribosome profiling, the deep sequencing of ribosome protected mRNA fragments, is a method that monitors protein synthesis with scale, speed, and accuracy rivaling approaches for mRNA measurement. I applied this method to numerous time points through the yeast meiotic program in parallel with mRNA-seq and molecular staging to generate a rich atlas of meiotic events and gene expression and the first high-resolution map of protein synthesis through a developmental program (Brar et al., Science, 2012).
This study revealed an unprecedented view of the molecular events underlying diverse aspects of meiotic biology and uncovered numerous new and dramatic instances of dynamic translational regulation through meiosis. The effort also yielded several fundamental surprises with broad significance. Projects in the Brar lab are based on exploring the basis and molecular significance of these discoveries.
What is the role of the numerous short peptides translated in meiotic cells?
The above study uncovered surprising complexity to genome coding in meiotic yeast cells, including the translation of thousands of discrete regions between AUG codons and stop codons that were not previously thought to house genes. We systematically annotated these regions through a sensitive, empirical strategy that defined many novel Open Reading Frames (ORFs) on stable transcriptsas well as alternate isoforms of well characterized proteins, As nearly all of the new ORFs are short (sORFs), they have been excluded from traditional approaches requiring 80-100 amino acids for annotation, and cellular roles thus remain unexplored. Recent studies in diverse organisms have identified cellular functions for short peptides, although the function of the meiotic sORFs remains a mystery. The presence of the corresponding short peptide products specifically in meiotic cells can be verified by classical methods and these early studies suggest diverse function and a rich pool of factors for functional discovery.
What are the roles and mechanisms of induction for the UPR and other stress responsive pathways in meiosis?
Through meiotic differentiation, in the absence of dynamic external stimuli, timed induction of prominent stress pathway factors was observed in our original study. Many of these pathways include post-translational regulation, suggesting that these measurements underestimate their dynamic control. Nonetheless, the striking and stereotypical regulation of stress programs in meiosis suggests that while harsh exogenous treatment including heat and drugs may have enabled their discovery, categorization of such pathways as 'stress-responsive' may not reflect their sole or even major physiological role.
The DNA damage response is a case in which a conserved and dedicated role in meiotic differentiation is clear. In fact, studies of meiotic recombination have revealed general principles of DNA repair. I propose that this case reflects a broad theme. For instance, while central components of the Unfolded Protein Response (UPR) are conserved from yeast to human,they are dispensable for growth in yeast. Thus, study of the yeast UPR has relied on non-physiological experimentation, including the use of inhibitors of ER folding such as dithiothreitol (DTT). Drug-based studies have yielded significant insight into UPR function, but meiotic UPR induction provides the first physiological yeast model of timed induction akin to that seen in differentiating mammalian cells.
What are the factors responsible for the pervasive and dramatic translational regulation observed in cells undergoing meiosis?
The diversity of cellular changes within a meiotic cell is reflected by the large fraction of known genes (6134 out of 6708) that are translated in meiosis. Most show strong temporal regulation, with 66% of genes varying by at least 10-fold in expression within the meiotic program. Parallel measurements of mRNA and new translation provided the first glimpse of global translational control through a developmental process. This study showed that translational regulation contributes strongly to temporal tuning of gene expression through meiosis, with 24% of genes subject to strong translational control during meiosis. This dataset provides a valuable tool to probe the molecular basis for, and cellular importance of, such dynamic control and the identification of the cis- and trans-factors responsible.
What is different about translation in meiosis that results in use of many upstream Open Reading Frames (uORFs)?
Many instances of upstream open reading frame (uORFs) translation were observed in cells undergoing meiosis, suggesting a meiotic shift in translation mechanism for at least a subset of transcripts. Additionally, many of these uORFs initiate translation at near-cognate (non-AUG) codons. The uORFs observed are highly discrete, specific, and heavily enriched in meiosis relative to any other condition examined to date, but the molecular basis for these unusual observations remains a mystery. We aim to unravel both the biological significance of these uORFs as well as the translation specializations that produce them in a meiosis-specific manner.
Eisenberg AR, Higdon AL, Hollerer I, Fields AP, Jungreis I, Diamond PD, Kellis M, Jovanovic M, Brar GA.Translation Initiation Site Profiling Reveals Widespread Synthesis of Non-AUG-Initiated Protein Isoforms in Yeast. Cell Systems.2020.
Cheng Z and Brar GA. Global translation inhibition yields condition-dependent de-repression of ribosome biogenesis mRNAs. Nucleic Acids Research. 2019.
Cheng Z, Mugler CF, Keskin A, Hodapp S, Chan LY, Weis K, Mertins P, Regev A, Jovanovic M, Brar GA. Small and Large Ribosomal Subunit Deficiencies Lead to Distinct Gene Expression Signatures that Reflect Cellular Growth Rate. Molecular Cell. 2019.
Eisenberg AR, Higdon A, Keskin A, Hodapp S, Jovanovic M, Brar GA. Precise Post-translational Tuning Occurs for Most Protein Complex Components during Meiosis. Cell Reports. 2018.
Van Dalfsen KM, Hodapp S, Keskin A, Otto GM, Berdan CA, Higdon A, Cheunkarndee T, Nomura DK, Jovanovic M, Brar GA. Global Proteome Remodeling during ER Stress Involves Hac1-Driven Expression of Long Undecoded Transcript Isoforms. Developmental Cell. 2018.
Otto GM and Brar GA. Seq-ing answers: uncovering the unexpected in global gene regulation. Curr Genetics. 2018
Hollerer I, Barker JC, Jorgenson V, Tresenrider A, Dugast-Darzacq C, Chan LY, Darzacq X, Tjian R, Ünal, E*, Brar GA*. Evidence for an Integrated Gene Repression Mechanism based on mRNA Isoform Toggling in Human Cells biorxiv.org/content/early/2018/10/05/264721 (*equal contributions)
Guenther U-P, Weinberg DE, Zubradt MM, Tedeschi FA, Stawicki BN, Zagore LL, Brar GA, Licatalosi DD, Bartel DP, Weissman JS & Jankowsky E. The helicase Ded1p controls use of near-cognate translation initiation codons in 5′ UTRs. Nature. 2018.
Cheng Z*, Otto GM*, Powers EN, Keskin A, Mertins P, Carr SA, Jovanovic M, Brar GA. Pervasive, coordinated protein level changes driven by transcript isoform switching during meiosis. Cell. 2018 (*equal contributions)
Powers EN, Brar GA. m6A and eIF2α-p Team Up to Tackle ATF4 Translation during Stress. Mol Cell. 2018
Hollerer I, Higdon A, Brar GA. Strategies and Challenges in Identifying Function for Thousands of sORF-Encoded Peptides in Meiosis. Proteomics. 2017
Brar GA. Beyond the Triplet Code: Context Cues Transform Translation.Cell. Review. 2016 Dec 15;167(7):1681-1692
Brar GA and Weissman JS. Ribosome profiling reveals the what, when, where, and how of protein synthesis. Nature Reviews Molecular and Cell Biology. 2015 Nov;16(11):651-64.
Brar GA, Yassour M, Friedman N, Regev A, Ingolia NT, Weissman JS. High-Resolution View of the Yeast Meiotic Program Revealed by Ribosome Profiling. Science. Article. 2012 Feb 3;335(6068):552-7.
Ingolia NT, Brar GA, Stern-Ginossar N, Harris N, Talhouarne GJS, Jackson SE, Wills MR, and Weissman JS. Ribosome Profiling Reveals Pervasive Translation Outside of Annotated Protein-Coding Genes. Cell Reports (In Press).
Berchowitz LE, Gajadhar AS, van Werven FJ, De Rosa AA, Samoylova ML, Brar GA, Xu Y, Xiao C, Futcher B, Weissman JS, White FM, Amon A. A developmentally regulated translational control pathway establishes the meiotic chromosome segregation pattern. Genes Dev. 2013 Oct 1. PMID: 24115771.
Gilbert LA, Larson MH, Morsut L, Liu Z, Brar GA, Torres SE, Stern-Ginossar N, Onn Brandman, Whitehead EH, Doudna JA, Lim WA, Weissman JS, Qi LS. CRISPR-Mediated Modular RNA-Guided Regulation of Transcription in Eukaryotes. Cell. In press.
Thorburn RR, Gonzalez C, Brar GA, Christen S, Carlile TM, Ingolia NT, Sauer U, Weissman JS, Amon A. Aneuploid yeast strains exhibit defects in cell growth and passage through START. Mol Biol Cell. 2013 May; 24(9):1274-89.
Miller, M.P.*, Ünal, E.*, Brar, GA, Amon, A. (*equal contributions) Meiosis I chromosome segregation is established by inhibiting microtubule-kinetochore interactions in Prophase I. eLife. 2012 Dec 18; 1:e00117.
Ingolia NT, Brar GA, Rouskin S, McGeachy AM, Weissman JS. The ribosome profiling strategy for monitoring translation in vivo by deep sequencing of ribosome-protected mRNA fragments. Nature Protocols. 2012 Jul 26;7(8):1534-50.
Carvunis AR, Rolland T, Wapinski I, Calderwood MA, Yildirim MA, Simonis N, Charloteaux B, Hidalgo CA, Barbette J, Santhanam B, Brar GA, Weissman JS, Regev A, Thierry-Mieg N, Cusick ME, Vidal M. Proto-genes and de novo gene birth. Nature. 2012 Jul 19;487(7407):370-4.
Brar GA, Hochwagen A, Ee L, Amon A. The multiple roles of cohesin in meiotic chromosome morphogenesis and pairing. Mol Biol Cell. 2009 Feb;20(3):1030-47.
Brar GA and Amon A. Emerging roles for centromeres in meiosis I chromosome segregation. Nature Reviews Genetics. Review. 2008 Dec;9(12):899-910.
Brar GA, Kiburz BM, Zhang Y, Kim JE, White F, Amon A. Rec8 phosphorylation and recombination promote the step-wise loss of cohesins in meiosis. Nature. 2006 May 25;441(7092):532-6.
Hochwagen A, Tham WH, Brar GA, Amon A. The FK506 binding protein Fpr3 counteracts protein phosphatase 1 to maintain meiotic recombination checkpoint activity. Cell. 2005 Sep 23;122(6):861-73.
Photo credit: Mark Hanson at Mark Joseph Studios.
Last Updated 2020-10-01