Project description:PA1 has been identified as a component of a MLL3/4-containing histone methyltransferase complex. PA1 directly interacts with PTIP but not with other complex components. Since biological functions of PA1 are unknown, we used microarrays to determine which genes are regulated by PA1. To identify PA1-regulated genes, immortalized PA1 conditional knockout PA1loxP/loxP MEFs were infected with retroviruses expressing either Cre recombinase or vector alone. We prepared duplicated RNAs from either vector or Cre infected cells (PA1+/+ or PA1-/-) for 4 affymetrix microarrays.
Project description:PA1 has been identified as a component of a MLL3/4-containing histone methyltransferase complex. PA1 directly interacts with PTIP but not with other complex components. Since biological functions of PA1 are unknown, we used microarrays to determine which genes are regulated by PA1.
Project description:Blood-testis barrier (BTB) is essential to the microenvironment of spermatogenesis, and Sertoli cells provide the cellular basis for BTB construction. Numerous nuclear transcription factors have been identified to be vital for the proper functions of Sertoli cells. PA1 has been reported to play important roles during diverse biological processes, while its potential function in male reproduction is still unknown. Here, we show that PA1 was highly expressed in human and mouse testis and predominantly localized in the nuclei of Sertoli cells. Sertoli cell-specific Pa1 knockout resulted in azoospermia-like phenotype in mice. The knockout of this gene lead to multiple defects in spermatogenesis, such as the disorganization of cytoskeleton at basal and apical ectoplasmic specialization and the disruption of the BTB. Further transcriptomic analysis together with Cut-Tag results of PA1 in Sertoli cells revealed that PA1 could affect the expression of a subset of genes which are essential for the normal functions of Sertoli cells including those genes associated with actin organization and cellular junction such as Connexin43 (Cx43). We further demonstrated that the expression of Cx43 depended on the interaction between JUN, one of the AP-1 complex transcription factors, and PA1. Overall, our findings firstly revealed that PA1 is essential to the maintaining of BTB integrity in Sertoli cells, it regulates BTB construction-related gene expression via interacting a transcription factor, thus providing a potential diagnostic or even therapeutic target for some azoospermia patients.
Project description:This study profiles RNA:DNA hybrid formation in human and mouse cell lines. DRIPc-seq (strand-specific R-loop mapping) was performed on human NT2 cells and mouse 3T3 cells. DRIP-seq (R-loop mapping) was performed on human NT2 and K562 and mouse E14 and 3T3 cell lines. MethylC-seq and RNA-seq were performed on NT2.
