Changes in circulating levels of immunoreactive follicle stimulating hormone, luteinizing hormone, and testosterone during sexual development in the rhesus monkey, Macaca mulatta

Basal serum levels of follicle stimulating hormone (FSH), luteinizing hormone (LH), and testosterone (T) and the responsiveness of these hormones to a challenge dose of luteinizing hormone releasing hormone (LHRH), were determined in juvenile, pubertal, and adult rhesus monkeys. The monkey gonadotrophins were analyzed using RIA reagents supplied by the World Health Organization (WHO) Special Programme of Human Reproduction. The FSH levels which were near the assay sensitivity in immature monkeys (2.4 +/- 0.8 ng/ml) showed a discernible increase in pubertal animals (6.4 +/- 1.8 ng/ml). Compared to other two age groups, the serum FSH concentration was markedly higher (16.1 +/- 1.8 ng/ml) in adults. Serum LH levels were below the detectable limits of the assay in juvenile monkeys but rose to 16.2 +/- 3.1 ng/ml in pubertal animals. When compared to pubertal animals, a two-fold increase in LH levels paralleled changes in serum LH during the three developmental stages. Response of serum gonadotrophins and T levels to a challenge dose of LHRH (2.5 micrograms; i.v.) was variable in the different age groups. The present data suggest: an asynchronous rise of FSH and LH during the pubertal period and a temporal correlation between the testicular size and FSH concentrations; the challenge dose of LHRH, which induces a significant rise in serum LH and T levels, fails to elicit an FSH response in all the three age groups; and the pubertal as compared to adult monkeys release significantly larger quantities of LH in response to exogenous LHRH.     

Profiles of gonadotropins and steroid hormone-like substances in the hemolymph of mud crab Scylla paramamosain during the reproduction cycle

To elucidate the role of gonadotropins-like substances in mud crab Scylla paramamosain, hemolymph samples were measured for concentrations of follicle stimulating hormone (FSH), luteinizing hormone (LH) and steroid hormones by enzyme-linked immunosorbent assay (ELISA). Hormonal concentration data were analyzed in association with the stages of gonadal development. ELISA has shown that in the female crab, the level of FSH reaches its peak in the early stage of ovary development, while estradiol and LH peaked during the late maturing stage of the ovary. In the male crab, testosterone and FSH culminated during the spermatid stage, and the level of LH peaked during the sperm stage. These results indicated that substances resembling the vertebrate FSH and LH are present in the hemolymph of S. paramamosain, and they may be involved in the development of the gonad.     

Expression and regulation of protein kinase CK2 during the cell cycle

There are indications from genetic, biochemical and cell biological studies that protein kinase CK2 (formerly casein kinase II) has a variety of functions at different stages in the cell cycle. To further characterize CK2 and its potential roles during cell cycle progression, one of the objectives of this study was to systematically examine the expression of all three subunits of CK2 at different stages in the cell cycle. To achieve this objective, we examined levels of CK2, CK2 and CK2 on immunoblots as well as CK2 activity in samples prepared from: (i) elutriated populations of MANCA (Burkitt lymphoma) cells, (ii) serum-stimulated GL30-92/R (primary human fibroblasts) cells and (iii) drug-arrested chicken bursal lymphoma BK3A cells. On immunoblots, we observed a significant and co-ordinate increase in the expression of CK2 and CK2 following serum stimulation of quiescent human fibroblasts. By comparison, no major fluctuations in CK2 activity were detected during any other stages during the cell cycle. Furthermore, we did not observe any dramatic differences between the relative levels of CK2 to CK2 during different stages in the cell cycle. However, we observed a significant increase in the amount of CK2 relative to CK2 in cells arrested with nocodazole. We also examined the activity of CK2 in extracts or in immunoprecipitates prepared from drug-arrested cells. Of particular interest is the observation that the activity of CK2 is not changed in nocodazole-arrested cells. Since CK2 is maximally phosphorylated in these cells, this result suggests that the phosphorylation of CK2 by p34cdc2 does not affect the catalytic activity of CK2. However, the activity of CK2 was increased by incubation with p34cdc2 in vitro. Since this activation was independent of ATP we speculate that p34cdc2 may have an associated factor that stimulates CK2 activity. Collectively, the observations that relative levels of CK2 increase in mitotic cells, that CK2 and CK2 are phosphorylated in mitotic cells and that p34cdc2 affects CK2 activity in vitro suggest that CK2 does have regulatory functions associated with cell division.     

oca2 regulation of chromatophore differentiation and number is cell type specific in zebrafish

We characterized a zebrafish mutant that displays defects in melanin synthesis and in the differentiation of melanophores and iridophores of the skin and retinal pigment epithelium. Positional cloning and candidate gene sequencing link this mutation to a 410‐kb region on chromosome 6, containing the oculocutaneous albinism 2 (oca2) gene. Quantification of oca2 mutant melanophores shows a reduction in the number of differentiated melanophores compared with wildtype siblings. Consistent with the analysis of mouse Oca2‐deficient melanocytes, zebrafish mutant melanophores have immature melanosomes which are partially rescued following treatment with vacuolar‐type ATPase inhibitor/cytoplasmic pH modifier, bafilomycin A1. Melanophore‐specific gene expression is detected at the correct time and in anticipated locations. While oca2 zebrafish display unpigmented gaps on the head region of mutants 3 days post‐fertilization, melanoblast quantification indicates that oca2 mutants have the correct number of melanoblasts, suggesting a differentiation defect explains the reduced melanophore number. Unlike melanophores, which are reduced in number in oca2 mutants, differentiated iridophores are present at significantly higher numbers. These data suggest distinct mechanisms for oca2 in establishing differentiated chromatophore number in developing zebrafish.     

Cell recognition molecule L1 promotes embryonic stem cell differentiation through the regulation of cell surface glycosylation

Cell recognition molecule L1 (CD171) plays an important role in neuronal survival, migration, differentiation, neurite outgrowth, myelination, synaptic plasticity and regeneration after injury. Our previous study has demonstrated that overexpressing L1 enhances cell survival and proliferation of mouse embryonic stem cells (ESCs) through promoting the expression of FUT9 and ST3Gal4, which upregulates cell surface sialylation and fucosylation. In the present study, we examined whether sialylation and fucosylation are involved in ESC differentiation through L1 signaling. RNA interference analysis showed that L1 enhanced differentiation of ESCs into neurons through the upregulation of FUT9 and ST3Gal4. Furthermore, blocking the phospholipase Cγ (PLCγ) signaling pathway with either a specific PLCγ inhibitor or knockdown PLCγ reduced the expression levels of both FUT9 and ST3Gal4 mRNAs and inhibited L1-mediated neuronal differentiation. These results demonstrate that L1 promotes neuronal differentiation from ESCs through the L1-mediated enhancement of FUT9 and ST3Gal4 expression.     

Macrophage migration inhibitory factor in the regulation of myoblast proliferation and differentiation

Obesity is documented to be a state of chronic mild inflammation associated with increased macrophage infiltration into adipose tissue and liver and skeletal muscle. As a pleiotropic inflammatory mediator, macrophage migration inhibitory factor (MIF) is associated with metabolic disease, so MIF may signal molecular links between adipocytes and myocytes. MIF expression was modified during myoblast differentiation, but the role of MIF during this process is unclear. C2C12 cells were transfected with MIF to investigate their role during differentiation. MIF expression attenuated C2C12 differentiation. It did not change proliferation, but downregulated cyclin D1 and CDK4, causing cell accumulation in the G1 phase. p21 protein was increased significantly and MyoD, MyoG, and p21 mRNA also increased significantly in the C2C12 cells treated with ISO-1, suggesting that inhibition of MIF promotes differentiation. MIF inhibits the myoblast differentiation by affecting the cell cycle progression, but does not affect proliferation.     

A novel protein,sperm head and tail associated protein (SHTAP), interacts with cysteine‐rich secretory protein 2 (CRISP2) during spermatogenesis in the mouse

Background information. CRISP2 (cysteine‐rich secretory protein 2) is a sperm acrosome and tail protein with the ability to regulate Ca²⁺ flow through ryanodine receptors. Based on these properties, CRISP2 has a potential role in fertilization through the regulation of ion signalling in the acrosome reaction and sperm motility. The purpose of the present study was to determine the expression, subcellular localization and the role in spermatogenesis of a novel CRISP2‐binding partner, which we have designated SHTAP (sperm head and tail associated protein). Results. Using yeast two‐hybrid screens of an adult testis expression library, we identified SHTAP as a novel mouse CRISP2‐binding partner. Sequence analysis of all Shtap cDNA clones revealed that the mouse Shtap gene is embedded within a gene encoding the unrelated protein NSUN4 (NOL1/NOP2/Sun domain family member 4). Five orthologues of the Shtap gene have been annotated in public databases. SHTAP and its orthologues showed no significant sequence similarity to any known protein or functional motifs, including NSUN4. Using an SHTAP antiserum, multiple SHTAP isoforms (～20–87 kDa) were detected in the testis, sperm, and various somatic tissues. Interestingly, only the ～26 kDa isoform of SHTAP was able to interact with CRISP2. Furthermore, yeast two‐hybrid assays showed that both the CAP (CRISP/antigen 5/pathogenesis related‐1) and CRISP domains of CRISP2 were required for maximal binding to SHTAP. SHTAP protein was localized to the peri‐acrosomal region of round spermatids, and the head and tail of the elongated spermatids and sperm tail where it co‐localized with CRISP2. During sperm capacitation, SHTAP and the SHTAP—CRISP2 complex appeared to be redistributed within the head. Conclusions. The present study is the first report of the identification, annotation and expression analysis of the mouse Shtap gene. The redistribution observed during sperm capacitation raises the possibility that SHTAP and the SHTAP—CRISP2 complex play a role in the attainment of sperm functional competence.     

细胞终末分化过程中三维基因组结构与功能调控的分子机制

真核细胞中的染色质DNA高度折叠形成复杂的三维结构,其空间组织方式对精准调控基因的表达和细胞发挥正常功能都起着重要的作用。细胞终末分化成熟过程中形态及基因表达谱常发生显著改变,同时伴随着明显的基因组三维结构变化。本文在简单介绍三维基因组多层次组织结构(染色质领域、A/B区室、拓扑相关结构域和成环构象等)基础上,重点综述了细胞终末分化过程中三维基因组结构变化与功能调控方面的研究进展,并探讨了当前三维基因组研究在解析细胞分化成熟过程时存在的问题和前景。