Mouse TMCO5 is localized to the manchette microtubules involved in vesicle transfer in the elongating spermatids.

As a result of a high-throughput in situ hybridization screening for adult mouse testes, we found that the mRNA for Tmco5 is expressed in round and elongating spermatids. Tmco5 belongs to the Tmco (Transmembrane and coiled-coil domains) gene family and has a coiled-coil domain in the N-terminal and a transmembrane domain in the C-terminal region. A monoclonal antibody raised against recombinant TMCO5 revealed that the protein is expressed exclusively in the elongating spermatids of step 9 to 12 and is localized to the manchette, a transiently emerging construction, which predominantly consists of cytoskeleton microtubules and actin filaments.

Xenopus Vasa Homolog XVLG1 is Essential for Migration and Survival of Primordial Germ Cells.

Xenopus vasa-like gene 1 (XVLG1), a DEAD-Box Helicase 4 (DDX4) gene identified as a vertebrate vasa homologue, is required for the formation of primordial germ cells (PGCs). However, it remains to be clarified when and how XVLG1 functions in the formation of the germ cells. To gain a better understanding of the molecular mechanisms underlying XVLG1 during PGC development, we injected XVLG1 morpholino oligos into germ-plasm containing blastomeres of 32-cell stage of Xenopus embryos, and traced cell fates of the injected blastomere-derived PGCs.

Cell-mass structures expressing the aromatase gene Cyp19a1 lead to ovarian cavities in Xenopus laevis.

The African clawed frog, Xenopus laevis, has a ZZ/ZW-type sex-determination system. We previously reported that a W-linked gene, Dm-W, can determine development as a female. However, the mechanisms of early sex differentiation remain unclear.

Cytoglobin is expressed in hepatic stellate cells, but not in myofibroblasts, in normal and fibrotic human liver.

Cytoglobin (CYGB) is ubiquitously expressed in the cytoplasm of fibroblastic cells in many organs, including hepatic stellate cells. As yet, there is no specific marker with which to distinguish stellate cells from myofibroblasts in the human liver. To investigate whether CYGB can be utilized to distinguish hepatic stellate cells from myofibroblasts in normal and fibrotic human liver, human liver tissues damaged by infection with hepatitis C virus (HCV) and at different stages of fibrosis were obtained by liver biopsy.

Protein disulfide isomerase P5-immunopositive inclusions in patients with Alzheimer's disease.

Amyloid plaques and neurofibrillary tangles (NFTs) are the major pathological characteristics of Alzheimer's disease (AD). NFTs are composed of tubular filaments and paired helical filaments containing polymerized hyperphosphorylated tau protein. Another feature of AD is excessive generation of nitric oxide (NO).

Depression of mitochondrial metabolism by downregulation of cytoplasmic deacetylase, HDAC6.

Mitochondria perform multiple functions critical to the maintenance of cellular homeostasis. Here we report that the downregulation of histone deacetylase 6 (HDAC6) causes a reduction in the net activity of mitochondrial enzymes, including respiratory complex II and citrate synthase. HDAC6 deacetylase and ubiquitin-binding activities were both required for recovery of reduced mitochondrial metabolic activity due to the loss of HDAC6.

Mitochondrial P5, a member of protein disulphide isomerase family, suppresses oxidative stress-induced cell death.

P5, one of the protein disulphide isomerase (PDI) family members, catalyses disulphide bond formation in proteins and exhibits molecular chaperone and calcium binding activities in vitro, whereas its physiological significance remains controversial. Recently, we have reported that P5 localizes not only in the ER but also in mitochondria, although it remains unclear so far about its physiological significance(s) of its dual localization. Here we report that H(2)O(2)- or rotenone-induced cell death is suppressed in MTS-P5 cells, which stably express P5 in mitochondria.

Characterization of the 38 kDa protein lacking in gastrula-arrested mutant Xenopus embryos.

We have reported elsewhere that offspring from the No. 65 female of Xenopus laevis cleaved normally, but their development was arrested at the onset of gastrulation, like the Ambystoma ova-deficient (o) mutant, irrespective of mating with different wild-type males, and that an acidic, 38 kDa protein present in wild-type eggs was lacking in eggs of the female. In the current study, we first determined the partial amino acid sequence (VANLE) of one of the well-separated tryptic peptides from the protein, which was found in elongation factor 1 delta (Ef1delta) in Xenopus, and finally identified the protein as one of the Ef1delta isoforms, Ef1delta2, by peptide mass spectrometry.

Analysis of SDF-1/CXCR4 signaling in primordial germ cell migration and survival or differentiation in Xenopus laevis.

Directional migration of primordial germ cells (PGCs) toward future gonads is a common feature in many animals. In zebrafish, mouse and chicken, SDF-1/CXCR4 chemokine signaling has been shown to have an important role in PGC migration. In Xenopus, SDF-1 is expressed in several regions in embryos including dorsal mesoderm, the target region that PGCs migrate to.

A novel method for microinjection into Xenopus eggs and embryos supported in methylcellulose solution.

We have developed a novel method for microinjection into Xenopus eggs and embryos. Microinjection was performed into eggs or embryos that were placed in wells (ca. 2.

Evidence for mitochondrial localization of P5, a member of the protein disulphide isomerase family.

This report demonstrates for the first time that P5, a member of the protein disulphide isomerase (PDI) family, is present in the mitochondria. Various organelles were screened for proteins bearing the CGHC motif using an affinity column conjugated with the phage antibody 5E, which cross-reacts with PDI family proteins. P5 was found in bovine liver mitochondrial extract and identified by Western blot analysis using anti-P5 antibody and by mass spectrometric analysis.

Ectopic germline cells in embryos of Xenopus laevis.

Whether all descendants of germline founder cells inheriting the germ plasm can migrate correctly to the genital ridges and differentiate into primordial germ cells (PGCs) at tadpole stage has not been elucidated in Xenopus. We investigated precisely the location of descendant cells, presumptive primordial germ cells (pPGCs) and PGCs, in embryos at stages 23-48 by whole-mount in situ hybridization with the antisense probe for Xpat RNA specific to pPGCs and whole-mount immunostaining with the 2L-13 antibody specific to Xenopus Vasa protein in PGCs. Small numbers of pPGCs and PGCs, which were positively stained with the probe and the antibody, respectively, were observed in ectopic locations in a significant number of embryos at those stages.

Methionine aminopeptidase II: A molecular chaperone for sarcoplasmic reticulum calcium ATPase.

The monoclonal antibody to the beta-subunit of H(+)/K(+)-ATPase (mAbHKbeta) cross-reacts with a protein that acts as a molecular chaperone for the structural maturation of sarcoplasmic reticulum (SR) Ca(2+)-ATPase. We partially purified a mAbHKbeta-reactive 65-kDa protein from Xenopus ovary. After in-gel digestion and peptide sequencing, the 65-kDa protein was identified as methionine aminopeptidase II (MetAP2).

The bHLH transcription factor Tcf12 (ME1) mRNA is abundantly expressed in Paneth cells of mouse intestine.

Using a large-scale in situ hybridization screening system, we found that mRNA coding for ME1, a basic helix-loop-helix (bHLH) transcription factor, was abundantly expressed in Paneth cells of adult small intestinal crypts. Other functionally related E-protein mRNAs, ME2, and E2A, however, could not be detected in the cells. ME1 mRNA was first detected in the jejunum and ileum two weeks after birth when the number of Paneth cells starts to increase.

The mRNA coding for Xenopus glutamate receptor interacting protein 2 (XGRIP2) is maternally transcribed, transported through the late pathway and localized to the germ plasm.

Using a large-scale in situ hybridization screening, we found that the mRNA coding for Xenopus glutamate receptor interacting protein 2 (XGRIP2) was localized to the germ plasm of Xenopus laevis. The mRNA is maternally transcribed in oocytes and, during maturation, transported to the vegetal germ plasm through the late pathway where VegT and Vg1 mRNAs are transported. In the 3'-untranslated region (UTR) of the mRNA, there are clusters of E2 and VM1 localization motifs that were reported to exist in the mRNAs classified as the late pathway group.

The Xdsg protein in presumptive primordial germ cells (pPGCs) is essential to their differentiation into PGCs in Xenopus.

In order to know the role of the Xdsg gene in presumptive PGCs (pPGCs) of Xenopus, we attempted to inhibit the translation of Xdsg mRNA in pPGCs by injecting antisense morpholino oligo (asMO), together with Fluorescein Dextran-Lysine (FDL), into single germ plasm-bearing cells of 32-cell embryos. Among three types of asMOs complementary to different parts of the 5'-untranslated region of Xdsg mRNA tested, only one asMO, designated as Xdsg-3, inhibited the translation of the mRNA in FDL-labeled pPGCs, resulting in the absence of labeled PGCs in experimental tadpoles. On the other hand, two other asMOs, Xdsg-1 and -2, did not inhibit the translation, so that a similar number of labeled PGCs found in FDL-injected but asMO-uninjected control tadpoles were observed in experimental tadpoles derived from asMO-injected embryos.

The mode and molecular mechanisms of the migration of presumptive PGC in the endoderm cell mass of Xenopus embryos.

We investigated the mode of migration of presumptive primordial germ cells (pPGC) in the endoderm cell mass of Xenopus embryos at stages 7-40. The molecules underlying the migration were also studied cytochemically and immunocytologically. By examining the relative positions of pPGC and somatic cells derived from the single, fluorescein-dextran lysine (FDL)-injected, germ plasm-bearing cells of stage 6 embryos, pPGC in embryos at stages 7-23 and those at stages later than 24 were assumed to passively and actively migrate in the endoderm cell mass, respectively.

Identification of novel keratinocyte-secreted peptides dermokine-alpha/-beta and a new stratified epithelium-secreted protein gene complex on human chromosome 19q13.1.

We performed high-throughput in situ hybridization screening of sections of mouse epidermis using an equalized skin cDNA library as probes and identified a novel gene giving rise to two splicing variants, both of which are expressed in the spinous layer. This gene was mapped between two genes encoding keratinocyte-related peptides, suprabasin and keratinocyte differentiation-associated protein (Kdap), on human chromosome 19q13.1.

Spatio-temporal distribution of the protein of Xenopus vasa homologue (Xenopus vasa-like gene 1, XVLG1) in embryos.

In order to know when the protein of Xenopus vasa homolog (Xenopus vasa-like gene 1, XVLG1) first appears in germ line cells and whether the protein is also present in somatic cells as is vasa protein in Drosophila, the spatio-temporal distribution of the protein in Xenopus embryos was carefully investigated by fluorescent microscopy. Part of the observation was performed by whole-mount immunocytochemistry and immunoblotting. A distinct fluorescence of XVLG1 protein was first recognized in a juxta-nuclear location of germ line cells or presumptive primordial germ cells (pPGC) at stage 12 (late gastrula) and remained associated with the pPGC or primordial germ cells (PGC) throughout the following stages until stage 46 (feeding tadpole).