The fate of pluripotent stem cells is tightly controlled during early embryonic development. Both the derivation and the maintenance of embryonic stem cells (ES cells) in vitro depend on feeder cell-derived growth factors that are largely unidentified. To dissect the mechanisms governing pluripotency, we conducted a screen to identify factors that are produced by mouse embryonic fibroblast STO cells and are required to maintain the pluripotency of ES cells. One of the factors is bone morphogenetic protein 4 (BMP4). Unexpectedly, the major effect of BMP4 on the self-renewal of ES cells is accomplished by means of the inhibition of both extracellular receptor kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) pathways, and inhibitors of ERK and p38 MAPKs mimic the effect of BMP4 on ES cells. Importantly, inhibition of the p38 MAPK pathway by SB203580 overcomes the block in deriving ES cells from blastocysts lacking a functional Alk3, the BMP type IA receptor. These results uncover a paradigm for BMP signaling in the biology of pluripotent stem cells.E mbryonic stem cells (ES cells) are able to form all cell types of the body by following normal embryogenesis (1-3). The pluripotency of ES cells has attracted great attention for their potential use in tissue and cell therapy. However, the molecular and developmental mechanisms controlling pluripotency and differentiation of ES cells are largely unknown, and only a very limited number of genes has so far been shown to affect the fate decisions of inner cell mass (ICM) or ES cells. These genes include Oct4, Fgf4, H2az, Foxd3, Nanog,.Growth factors required for ES cell self-renewal are usually provided by feeder cells, or exogenously (13). Leukemiainhibiting factor (LIF) and its close relatives are the known propluripotency factors for mouse ES cells. It is unclear how many other growth factors or signaling pathways are required for the self-renewal of ES cells. To address these questions, we set out to identify such factors that affect the self-renewal of ES cells. To accomplish this, we isolated and screened sublines of the mouse embryonic fibroblast STO cells for their ability to support the self-renewal of ES cells by using Oct4-GFP as a convenient marker of pluripotency (14-16). By this approach in combination with gene expression profiling, we have identified bone morphogenetic protein 4 (BMP4) as part of the extracellular propluripotency cues. Also, our studies show that BMP4 and LIF have synergistic effect in teratoma formation. Moreover, a number of genes differentially expressed in ES cells cultured with or without exogenous BMP4 have been identified.One of these differentially expressed genes, X-linked inhibitor of apoptosis (Xiap), is expressed at higher levels in ES cells cultured in the presence of exogenous BMP4 than in its absence. XIAP has been implicated in connecting the type I receptors of BMPs and TGF-␤s with the mitogen-activated protein kinase (MAPK) p38 pathway (17)(18)(19)(20). Contrary to previous findings in which BMP signaling up-r...

We previously identified a sperm-specific Na ؉ /H ؉ exchanger (sNHE) principally localized to the flagellum. Disruption of the sNHE gene in mice resulted in absolute male infertility associated with a complete loss of sperm motility. Here, we show that the sNHE-null spermatozoa fail to develop the cAMP-dependent protein tyrosine phosphorylation that coincides with the functional maturation occurring upon incubation in capacitating conditions in vitro. Both the sperm motility defect and the lack of induced protein tyrosine phosphorylation are rescued by the addition of cell-permeable cAMP analogs, suggesting that cAMP metabolism is impaired in spermatozoa lacking sNHE. Our analyses of the bicarbonate-dependent soluble adenylyl cyclase (sAC) signaling pathway in sNHE-null sperm cells reveal that sNHE is required for the expression of full-length sAC, and that it is important for the bicarbonate stimulation of sAC activity in spermatozoa. Furthermore, both codependent expression and coimmunoprecipitation experiments indicate that sNHE and sAC associate with each other. Thus, these two proteins appear to be components of a signaling complex at the sperm flagellar plasma membrane. We propose that the formation of this complex efficiently modulates intracellular pH and bicarbonate levels through the rapid and effective control of sAC and sNHE activities to facilitate sperm motility regulation.cAMP ͉ sperm motility

A family of integral membrane proteins containing a signature DHHC motif has been shown to display protein S-acyltransferase activity, modifying cysteine residues in proteins with fatty acids. The physiological roles of these proteins have largely been unexplored. Here we report that mice homozygous for a hypomorphic allele of a previously uncharacterized member, DHHC5, are born at half the expected rate, and survivors show a marked deficit in contextual fear conditioning, an indicator of defective hippocampal-dependent learning. DHHC5 is highly enriched in a post-synaptic density preparation and co-immunoprecipitates with post-synaptic density protein-95 (PSD-95), an interaction that is mediated through binding of the carboxyl terminus of DHHC5 and the PDZ3 domain of PSD-95. Immunohistochemistry demonstrated that DHHC5 is expressed in the CA3 and dentate gyrus in the hippocampus. These findings point to a previously unsuspected role for DHHC5 in post-synaptic function affecting learning and memory.Palmitoylation of proteins at intracellular sites is mediated by members of a family of S-fatty acyltransferases that are recognized by a DHHC consensus motif at the active site (1-4). In addition to functioning as protein S-acyltransferases, other functions have been proposed, with certain family members shown to transport divalent cations (5-6). The roles of DHHC proteins in the nervous system have attracted recent interest. Genetic alterations in DHHC9 and DHHC15 are responsible for two different forms of X-linked mental retardation in humans (7-9), whereas DHHC8 is contained within a microdeletion linked to schizophrenia in the Han Chinese population (10), and DHHC17 interacts with huntingtin (Htt) and may play a role in the pathogenesis of Huntington disease (11-13). Another DHHC family member, GODZ (DHHC3), has been shown to affect both excitatory and inhibitory synapse function (14 -19), and DHHC23 (neuronal nitric oxide synthetase (nNOS) 2 -interacting DHHC domain-containing protein with dendritic mRNA (NIDD)) targets neuronal nitric oxide synthetase to synapses via its nNOS PDZ binding domain (20). Given the large number of family members (22 in human and 23 in mouse) and the daunting number of potential substrates (over 250 in the brain, for example) (21), mouse models will become increasingly important for understanding their physiological roles. Of note, the only previously available DHHC gene knock-out mouse model (the DHHC8 model (22)), exhibits behavioral abnormalities and defects in neuronal culture (such as diminished density of dendritic spines) (23).In this study we show that a previously uncharacterized DHHC family member, DHHC5, is highly enriched in the brain and in synaptic vesicle fractions and that it interacts with the third PDZ binding domain of PSD-95 in vitro and in vivo. Mice homozygous for a hypomorphic allele of DHHC5 show impaired contextual fear conditioning. DHHC5 is shown to contain a hydroxylamine-sensitive acyl group in vivo, but PSD-95 palmitoylation is unchanged in DHHC5 knock-o...

This study reported the clonal dissemination of OXA-232-producing sequence type 15 (ST15) carbapenem-resistant Klebsiella pneumoniae among elderly patients in China. All patients were immunocompromised, suffered from multiple underlying diseases, and were hospitalized for a prolonged period; however, they slowly recovered on antimicrobial therapy. The blaOXA-232 gene was in a 6.1-kb ColKP3-type nonconjugative plasmid. The strains displayed a multidrug resistance phenotype and were not hypervirulent despite harboring a virulence plasmid. Active surveillance should be enforced to control further transmission.

Bone morphogenetic proteins (BMPs) play essential roles in many aspects of developmental biology. We have previously shown that Bmp7, Bmp8a, and Bmp8b of the 60A class of Bmp genes have additive effects in spermatogenesis and in maintaining the epididymal integrity of the caput and caudal regions. Here we report that Bmp4 of the Dpp class has a unique expression pattern in the developing testis and epididymis. Bmp4 heterozygous males on a largely C57BL/6 background show compromised fertility due to degeneration of germ cells, reduced sperm counts, and decreased sperm motility. More interestingly, some of these males show extensive degeneration of the epididymal epithelium in the corpus region, rather than in the caput and cauda regions as for Bmp7 and Bmp8 mutants. Thus, these genetic data reveal a region-specific requirement of different classes of BMPs for epididymal epithelium to survive and have significant implications on male reproductive health and perhaps birth control.