Acceptedfor publicationMay 19

ABSTRACT We and other investigators obtained evidence that platelets stimulate endothelin-i (ET-i) production at both message and protein levels in vascular endothelial cells (EC5),and that plate let-derived transforming growth factor-f31 (TGF-j3i) is respon sible for this stimulation. In the present study, we examined the effects of acidification or heat treatment, known to activate latent TGF-f31, on the platelet supernatant-induced Er-i pro duction in cultured porcine aortic ECs. Supematant of platelets (6.0 x 1O@ platelets/mI) aggregated by adenosine diphosphate contained large amounts of TGF-pi, but were almost in a latent form, and the proportion of active TGF-j31 in the supematant was increased markedly in the case of acidification or heat treatment. These treatments also significantly potentiated the supematant-induced stimulation of prepro Er-i mRNA expres sion and the Er-i release in ECs. Purified TGF-@i also en hanced Er-i release, dose-dependently, but the enhancement declined at the higher concentrations. Thus, powerful stimula tion of Er-i production by platelet supematant after acidifica tion or heat treatment cannot be explained only by increments in active TGF-131. The supematant-induced stimulation of Er-i synthesis was significantly inhibited by concomitant treatment of TGF-@i neutralizing antibody, but this inhibition was incom plete even at a concentration that abolished TGF-31 -induced maximal stimulation. These resutts suggest that platelet-induced stimulation and subsequent acidification and heat treatment induced potentiation on endothelial Er-i production depend closely on release and activation of TGF-@i derived from plate lets. However, when TGF-fii concentration is increased, this peptide may further stimulate Er-i production, probably through interactions with other platelet-derived substances

Meiotic cohesins mediate initial loading of HORMAD1 to the chromosomes and coordinate SC formation during meiotic prophase.

During meiotic prophase, sister chromatids are organized into axial element (AE), which underlies the structural framework for the meiotic events such as meiotic recombination and homolog synapsis. HORMA domain-containing proteins (HORMADs) localize along AE and play critical roles in the regulation of those meiotic events. Organization of AE is attributed to two groups of proteins: meiotic cohesins REC8 and RAD21L; and AE components SYCP2 and SYCP3. It has been elusive how these chromosome structural proteins contribute to the chromatin loading of HORMADs prior to AE formation. Here we newly generated Sycp2 null mice and showed that initial chromatin loading of HORMAD1 was mediated by meiotic cohesins prior to AE formation. HORMAD1 interacted not only with the AE components SYCP2 and SYCP3 but also with meiotic cohesins. Notably, HORMAD1 interacted with meiotic cohesins even in Sycp2-KO, and localized along cohesin axial cores independently of the AE components SYCP2 and SYCP3. Hormad1/Rad21L-double knockout (dKO) showed more severe defects in the formation of synaptonemal complex (SC) compared to Hormad1-KO or Rad21L-KO. Intriguingly, Hormad1/Rec8-dKO but not Hormad1/Rad21L-dKO showed precocious separation of sister chromatid axis. These findings suggest that meiotic cohesins REC8 and RAD21L mediate chromatin loading and the mode of action of HORMAD1 for synapsis during early meiotic prophase

Meiosis-specific ZFP541 repressor complex promotes developmental progression of meiotic prophase towards completion during mouse spermatogenesis

During spermatogenesis, meiosis is accompanied by a robust alteration in gene expression and chromatin status. However, it remains elusive how the meiotic transcriptional program is established to ensure completion of meiotic prophase. Here, we identify a protein complex that consists of germ-cell-specific zinc-finger protein ZFP541 and its interactor KCTD19 as the key transcriptional regulators in mouse meiotic prophase progression. Our genetic study shows that ZFP541 and KCTD19 are co-expressed from pachytene onward and play an essential role in the completion of the meiotic prophase program in the testis. Furthermore, our ChIP-seq and transcriptome analyses identify that ZFP541 binds to and suppresses a broad range of genes whose function is associated with biological processes of transcriptional regulation and covalent chromatin modification. The present study demonstrates that a germ-cell specific complex that contains ZFP541 and KCTD19 promotes the progression of meiotic prophase towards completion in male mice, and triggers the reconstruction of the transcriptional network and chromatin organization leading to post-meiotic development