The bromodomain is an evolutionarily conserved motif that binds acetylated lysines in histones and other proteins. The BET sub-class of mammalian bromodomain-containing proteins is unique in that its members contain two bromodomains (BD1 and BD2) and an extra terminal (ET) domain. There are four BET family members in mouse (and human) - Brd2, Brd3, Brd4, and Brdt - and they are expressed in a striking and dynamic pattern in the male germ line. We have generated a mutation in the mouse Brdt gene, designated Brdt BD1, that yields a truncated protein lacking the first of the two bromodomains. Homozygous Brdt BD1 progeny are viable but the males are sterile, producing fewer sperm that are morphologically abnormal.
Aim 1 will test the hypotheses that i) Brdt functions as part of a transcription complex that regulates a set of genes whose expression is essential for spermatogenesis, and ii) that the BD1 of Brdt is required for this regulation. We will identify genes whose expression is changed in the absence of BD1 by microarray analysis and concomitantly, examine the chromatin modification status of the H1t promoter as a model for Brdt-complex binding regions.
Aim 2 will test the alternative, but not mutually exclusive, hypothesis that Brdt-containing complexes function to mark regions of the spermatid genome for subsequent recognition by complexes that are involved in the unique changes in chromatin structure during spermiogenesis. ChIP with anti-Brdt antibodies followed by genome-wide sequencing using the Solexa/Illumina 1 G technology will be used.
Aim 3 will test the hypotheses that first, the two bromodomains of Brdt have distinct functions in modulating transcription and/or chromatin re-modeling during spermiogenesis;second, that Brdt may function in several stages and processes of spermatogenesis, in addition to spermiogenesis;and third, that the Brdt BD1-mutant allele is a hypomorphic allele. We will generate a mutant allele producing Brdt protein containing BD1 but lacking BD2 (Aim 3a) and a mutant Brdt allele completely lacking functional protein (Aim 3b). We predict that the resulting phenotypes will overlap in part but will be distinct from the BD1-deficient mutant. Understanding the function of Brdt during spermatogenesis will provide a powerful developmental model system for elucidating the role of the BET genes during normal differentiation. Importantly, spermatogenesis is also a physiologically relevant system in which histone acetylation is clearly linked to chromatin remodeling.
Brdt is a member of a sub-family of bromodomain-containing proteins which have recently been shown to have essential functions in diverse basic cellular functions from DNA replication to transcription to chromatin remodeling. Our targeted mutational analysis has shown that deletion of the first of the two bromodomains in Brdt in the mouse model leads to male sterility, but the animals are otherwise viable and the females are fertile. Our studies will provide important insight into the potential mis- function of human BRDT in cases of unexplained (or idiopathic) infertility in men and may provide a new and novel target for male contraception.
|Wang, Li; Wolgemuth, Debra J (2016) BET Protein BRDT Complexes With HDAC1, PRMT5, and TRIM28 and Functions in Transcriptional Repression During Spermatogenesis. J Cell Biochem 117:1429-38|
|Garcia-Gutierrez, Pablo; Juarez-Vicente, Francisco; Wolgemuth, Debra J et al. (2014) Pleiotrophin antagonizes Brd2 during neuronal differentiation. J Cell Sci 127:2554-64|
|Chachua, T; Goletiani, C; Maglakelidze, G et al. (2014) Sex-specific behavioral traits in the Brd2 mouse model of juvenile myoclonic epilepsy. Genes Brain Behav 13:702-12|
|Berkovits, Binyamin D; Wolgemuth, Debra J (2013) The role of the double bromodomain-containing BET genes during mammalian spermatogenesis. Curr Top Dev Biol 102:293-326|
|Gaucher, Jonathan; Boussouar, Faycal; Montellier, Emilie et al. (2012) Bromodomain-dependent stage-specific male genome programming by Brdt. EMBO J 31:3809-20|
|Berkovits, Binyamin D; Wang, Li; Guarnieri, Paolo et al. (2012) The testis-specific double bromodomain-containing protein BRDT forms a complex with multiple spliceosome components and is required for mRNA splicing and 3'-UTR truncation in round spermatids. Nucleic Acids Res 40:7162-75|
|Berkovits, Binyamin D; Wolgemuth, Debra J (2011) The first bromodomain of the testis-specific double bromodomain protein Brdt is required for chromocenter organization that is modulated by genetic background. Dev Biol 360:358-68|
|Shang, Enyuan; Cui, Qingping; Wang, Xiangyuan et al. (2011) The bromodomain-containing gene BRD2 is regulated at transcription, splicing, and translation levels. J Cell Biochem 112:2784-93|
|Velisek, Libor; Shang, Enyuan; Veliskova, Jana et al. (2011) GABAergic neuron deficit as an idiopathic generalized epilepsy mechanism: the role of BRD2 haploinsufficiency in juvenile myoclonic epilepsy. PLoS One 6:e23656|
|Shang, Enyuan; Wang, Xiangyuan; Wen, Duancheng et al. (2009) Double bromodomain-containing gene Brd2 is essential for embryonic development in mouse. Dev Dyn 238:908-17|