Cell-specific occupancy of an extended repertoire of CREM and CREB binding loci in male germ cells

Abstract

BackgroundCREB and CREM are closely related factors that regulate transcription in response to various stress, metabolic and developmental signals. The CREMτ activator isoform is selectively expressed in haploid spermatids and plays an essential role in murine spermiogenesis.ResultsWe have used chromatin immunoprecipitation coupled to sequencing (ChIP-seq) to map CREM and CREB target loci in round spermatids from adult mouse testis and spermatogonia derived GC1-spg cells respectively. We identify more than 9000 genomic loci most of which are cell-specifically occupied. Despite the fact that round spermatids correspond to a highly specialised differentiated state, our results show that they have a remarkably accessible chromatin environment as CREM occupies more than 6700 target loci corresponding not only to the promoters of genes selectively expressed in spermiogenesis, but also of genes involved in functions specific to other cell types. The expression of only a small subset of these target genes are affected in the round spermatids of CREM knockout animals. We also identify a set of intergenic binding loci some of which are associated with H3K4 trimethylation and elongating RNA polymerase II suggesting the existence of novel CREB and CREM regulated transcripts.ConclusionsWe demonstrate that CREM and CREB occupy a large number of promoters in highly cell specific manner. This is the first study of CREM target promoters directly in a physiologically relevant tissue in vivo and represents the most comprehensive experimental analysis of CREB/CREM regulatory potential to date.

Retinoic Acid Receptors Recognize the Mouse Genome through Binding Elements with Diverse Spacing and Topology

“…ChIP and ChIP-seq-ChIP and ChIP-seq experiments were performed on chromatin from murine embryonic stem cells grown as embryoid bodies for 4 days in the absence of RA and then treated for 2 h with all-trans-retinoic acid as previously described (9,11,12). Further details are provided in the supplemental Experimental Procedures.…”

Interlocked loops trigger lineage specification and stable fates in the Drosophila nervous system

“…The chromatin IP (ChIP) was performed according to standard protocols 56,57 with modification. A quantity of 100 mg of embryos w 1118 stage 11 was homogenized with a Dounce homogenizer in 1 Â PBS, 0.4% formaldehyde and incubated 10 min at RT.…”

Unique Aspects of Transcription Regulation in Male Germ Cells

“…In agreement with this idea, ChIP-seq experiments show widespread occupancy by CREM of target genes that have no specific role in spermatogenesis, but whose promoters are active in haploid cells (Martianov et al 2010). The euchromatin of round spermatids therefore seems rather permissive for transcription factor binding which, together with high levels of the basal transcription machinery, may lead to the expression of a large set of genes and generally higher mRNA levels.…”

“…Diminished expression of these genes may account for the reduced sperm count and abnormalities seen in heterozygous CREM mutant mice, but do not readily explain the apoptosis. ChIP from adult testis coupled to high throughput sequencing shows that CREM occupies the promoters of more than 6000 target genes in haploid cells (Martianov et al 2010). Comparison with transcriptome data shows that only a small subset of these genes are deregulated following CREM inactivation.…”

“…Comparison with transcriptome data shows that only a small subset of these genes are deregulated following CREM inactivation. The deregulated genes include the antiapoptotic factor Bcl6b and genes involved in signal transduction and metabolic processes (Beissbarth et al 2003;Martianov et al 2010). It is rather the deregulation of these genes that accounts for the apoptotic phenotype seen on CREM inactivation.…”