MicroRNA (miRNA) and other types of small regulatory RNAs play a crucial role in the regulation of gene expression in eukaryotes. Several distinct classes of small regulatory RNAs have been ...discovered in recent years. To extend the repertoire of small regulatory RNAs characterized in chickens we used a deep sequencing approach developed by Solexa (now Illumina Inc.). We sequenced three small RNA libraries prepared from different developmental stages of the chicken embryo (days five, seven, and nine) to produce over 9.5 million short sequence reads. We developed a bioinformatics pipeline to distinguish authentic mature miRNA sequences from other classes of small RNAs and short RNA fragments represented in the sequencing data. Using this approach we detected almost all of the previously known chicken miRNAs and their respective miRNA* sequences. In addition we discovered 449 new chicken miRNAs including 88 miRNA candidates. Of these, 430 miRNAs appear to be specific to the avian lineage. Another six new miRNAs had evidence of evolutionary conservation in at least one vertebrate species outside of the bird lineage. The remaining 13 putative miRNAs appear to represent chicken orthologs of known vertebrate miRNAs. We discovered 39 additional putative miRNA candidates originating from miRNA generating intronic sequences known as mirtrons.
The maturation of oocytes external to the body is referred to IVM and is utilised by many different species. IVM bypasses traditional superovulation methods, where several hormonal injections are ...administered. Unfortunately, the pregnancy rates using IVM oocytes is less than those matured in vivo. This has impeded the uptake of the technology in ART laboratories. This review aims to explore the effects of multiple elements during IVM, discussing not only those which improve and impact on oocyte viability but those that can have deleterious effects on the oocyte.
VAMP/synaptobrevin-associated proteins (VAPs) contain an N-terminal major sperm protein domain (MSPd) that is associated with amyotrophic lateral sclerosis. VAPs have an intracellular housekeeping ...function, as well as an extracellular signaling function mediated by the secreted MSPd. Here we show that the
VAP homolog VPR-1 is essential for gonad development.
null mutants are maternal effect sterile due to arrested gonadogenesis following embryo hatching. Somatic gonadal precursor cells and germ cells fail to proliferate fully and complete their respective differentiation programs. Maternal or zygotic
expression is sufficient to induce gonadogenesis and fertility. Genetic mosaic and cell type-specific expression studies indicate that
activity is important in the nervous system, germ line and intestine. VPR-1 acts in parallel to Notch signaling, a key regulator of germline stem cell proliferation and differentiation. Neuronal
expression is sufficient for gonadogenesis induction during a limited time period shortly after hatching. These results support the model that the secreted VPR-1 MSPd acts at least in part on gonadal sheath cell precursors in L1 to early L2 stage hermaphrodites to permit gonadogenesis.
The major sperm protein domain (MSPd) has an extracellular signaling function implicated in amyotrophic lateral sclerosis. Secreted MSPds derived from the
VAPB homolog VPR-1 promote mitochondrial ...localization to actin-rich I-bands in body wall muscle. Here we show that the nervous system and germ line are key MSPd secretion tissues. MSPd signals are transduced through the CLR-1 Lar-like tyrosine phosphatase receptor. We show that CLR-1 is expressed throughout the muscle plasma membrane, where it is accessible to MSPd within the pseudocoelomic fluid. MSPd signaling is sufficient to remodel the muscle mitochondrial reticulum during adulthood. An RNAi suppressor screen identified survival of motor neuron 1 (SMN-1) as a downstream effector. SMN-1 acts in muscle, where it colocalizes at myofilaments with ARX-2, a component of the Arp2/3 actin-nucleation complex. Genetic studies suggest that SMN-1 promotes Arp2/3 activity important for localizing mitochondria to I-bands. Our results support the model that VAPB homologs are circulating hormones that pattern the striated muscle mitochondrial reticulum. This function is crucial in adults and requires SMN-1 in muscle, likely independent of its role in pre-mRNA splicing.
Understanding reproductive processes in parasitic nematodes has the potential to lead to the informed design of new anthelmintics and control strategies. Little is known, however, about the molecular ...mechanisms underlying sex determination, gametogenesis and reproductive physiology for most parasitic nematodes. Together with comparative analyses of data for the free-living nematode Caenorhabditis elegans , molecular investigations are beginning to provide insights into the processes involved in reproduction and development in parasitic nematodes. Here, we review recent developments, focusing on technological aspects and on molecules associated with sex-specific differences in adult nematodes.
Full-length genes representing different isoforms of the ubiquitin-conjugating enzyme UBC-2 were isolated from Oesophagostomum dentatum, cloned and sequenced. The alignment of their sequences ...(designated Od-ubc-2.1 to Od-ubc-2.3) revealed nucleotide variation at three positions within the predicted open reading frame of 444 bp. Substitutions were at positions 141 (A↔G), 142 (A↔G) and 296 (T↔C). Both former substitutions resulted in amino acid changes from a glycine residue to an arginine residue, whereas the latter resulted in a change from isoleucine to threonine. Comparison of predicted OD-UBC-2 with UBC-2 (protein) homologues/orthologues from 12 other species representing nematodes, Drosophila melanogaster, Saccharomyces cerevisiae, mice and humans revealed identities between species varying from 77 to 100% at the amino acid level, and motifs associated with protein conformation and function were identified. While the function of a representative ubc-2 gene from O. dentatum could not be established in C. elegans, it is likely to play a key role in the catabolism of proteins and in the development of O. dentatum.
A cDNA encoding a predicted small heat shock protein, HSP20, was isolated from the parasitic nematode
Haemonchus contortus. This cDNA encoded a predicted protein of 156 amino acids, which had high ...sequence identity with other nematode small heat shock proteins. Southern blot analysis of genomic DNA suggested that in
H. contortus HSP20 is encoded by a single copy gene. The HSP20 transcript and protein were expressed in the infective larvae (L3), early L4 and adult stages, but expression was not increased by heat shock treatment. In situ hybridisation analysis was used to localise expression of HSP20 mRNA in the adult parasite. Similar HSPs (heat shock protein) were detected by Western blotting in
Ancylostoma caninum,
Dictyocaulus viviparus, and
Toxocara canis, but not in
Trichostronglyus colubriformis. The conservation of HSP20 in several different nematode species may reflect its importance to parasites that require mammalian hosts as a part of their development.
Index Descriptors and Abbreviations: Haemonchus contortus; nematode; small heat shock protein; L3, infective larvae; xL3, exsheathed L3; eL4, early L4; EST, expressed sequence tag; HSP20, heat shock protein 20; sHSP, small heat shock protein
The VAPB/ALS8 major sperm protein domain (vMSP) is implicated in amyotrophic lateral sclerosis and spinal muscular atrophy, yet its function in the nervous system is not well understood. In ...Caenorhabditis elegans and Drosophila, the vMSP is cleaved from its transmembrane anchor and secreted in a cell type-specific fashion. We show that vMSPs secreted by neurons act on Lar-like protein-tyrosine phosphatase and Roundabout growth cone guidance receptors expressed in striated muscle. This signaling pathway promotes Arp2/3-dependent actin remodeling and mitochondrial localization to actin-rich muscle I-bands. C. elegans VAPB mutants have mitochondrial localization, morphology, mobility, and fission/fusion defects that are suppressed by Lar-like receptor or Arp2/3 inactivation. Hence, growth cone guidance receptor pathways that remodel the actin cytoskeleton have unanticipated effects on mitochondrial dynamics. We propose that neurons secrete vMSPs to promote striated muscle energy production and metabolism, in part through the regulation of mitochondrial localization and function.
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► Motor neurons secrete the VAPB/ALS8 major sperm protein domain (MSP) ► Secreted MSPs signal through Robo and Lar-like receptors in striated muscle ► MSP/Robo signaling inhibits Lar-mediated actin remodeling ► This pathway influences mitochondrial localization, fission/fusion, and respiration
Mutations in MSP domain proteins cause the neurodegenerative disorders amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA). Han et al. show that neurons secrete the MSP domain to control Robo/Lar signaling in muscle. Downstream effects on cortical actin influence muscle mitochondrial dynamics and function, consistent with ALS and SMA.
Mutations in VAPB/ALS8 are associated with amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA), two motor neuron diseases that often include alterations in energy metabolism. We ...have shown that C. elegans and Drosophila neurons secrete a cleavage product of VAPB, the N-terminal major sperm protein domain (vMSP). Secreted vMSPs signal through Roundabout and Lar-like receptors expressed on striated muscle. The muscle signaling pathway localizes mitochondria to myofilaments, alters their fission/fusion balance, and promotes energy production. Here, we show that neuronal loss of the C. elegans VAPB homolog triggers metabolic alterations that appear to compensate for muscle mitochondrial dysfunction. When vMSP levels drop, cytoskeletal or mitochondrial abnormalities in muscle induce elevated DAF-16, the Forkhead Box O (FoxO) homolog, transcription factor activity. DAF-16 promotes muscle triacylglycerol accumulation, increases ATP levels in adults, and extends lifespan, despite reduced muscle mitochondria electron transport chain activity. Finally, Vapb knock-out mice exhibit abnormal muscular triacylglycerol levels and FoxO target gene transcriptional responses to fasting and refeeding. Our data indicate that impaired vMSP signaling to striated muscle alters FoxO activity, which affects energy metabolism. Abnormalities in energy metabolism of ALS patients may thus constitute a compensatory mechanism counterbalancing skeletal muscle mitochondrial dysfunction.
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