Characterizing Variants of Unknown Significance of the PTEN tumour suppressorHomolog DAF-18.

Insulin and insulin-like growth factor signaling (IIS) is an anabolic pathway conserved among humans and Caenorhabditis elegans . In humans, the tumour suppressor protein Phosphatase and Tensin Homolog (PTEN) inhibits IIS, preventing excessive growth. PTEN variants are associated with disease, but how they affect PTEN function is not well understood. Here, we characterized variants of unknown significance (VUSs) implicated in autism spectrum disorder by studying homologous mutations in the C. elegans protein DAF-18 to infer how they play a role in human disease.We found that variants D66E and L115V are likely benign, H168Q is intermediate while variants H138R and T176I are likely pathogenic.

Transcriptomic and proteomic analyses of venom glands from scorpions Liocheles australasiae, Mesobuthus martensii, and Scorpio maurus palmatus.

Scorpion venom contains a cocktail of differing peptides and proteins. Previous studies focused on the identification of species-specific components in scorpion venoms, and whether there could be peptides and/or proteins conserved in the venom gland of a scorpion ancestor has been rarely investigated. Here, using a combination of transcriptomic and proteomic approaches, putative conserved toxins from the venom glands of scorpions Liocheles australasiae, Mesobuthus martensii, and Scorpio maurus palmatus were identified and compared. Similar to other studies, more than half of the conserved toxins are predominantly proteins including proteases. On the other hand, unique venom peptides, including ion channel toxins were revealed specifically in the M. martensii. The sodium channel toxin peptides revealed in M. martensii consolidated that scorpions in the Buthidae are able to envenomate their prey wih highly neurotoxic venom. This study suggested that these conserved proteins had already formed part of the arsenal in the venom gland of the common ancestor of scorpions, and likely perform important functional roles in envenomation during scorpion evolution.

An overview of the insulin signaling pathway in model organisms Drosophila melanogaster and Caenorhabditis elegans.

The insulin/insulin-like growth factor signaling pathway is an evolutionary conserved pathway across metazoans and is required for development, metabolism and behavior. This pathway is associated with various human metabolic disorders and cancers. Thus, model organisms including Drosophila melanogaster and Caenorhabditis elegans provide excellent opportunities to examine the structure and function of this pathway and its influence on cellular metabolism and proliferation. In this review, we will provide an overview of human insulin and the human insulin signaling pathway and explore the recent discoveries in model organisms Drosophila melanogaster and Caenorhabditis elegans. Our review will provide information regarding the various insulin-like peptides in model organisms as well as the conserved functions of insulin signaling pathways. Further investigation of the insulin signaling pathway in model organisms could provide a promising opportunity to develop novel therapies for various metabolic disorders and insulin-mediated cancers.

Horseshoe crab genomes reveal the evolution of genes and microRNAs after three rounds of whole genome duplication.

Whole genome duplication (WGD) has occurred in relatively few sexually reproducing invertebrates. Consequently, the WGD that occurred in the common ancestor of horseshoe crabs ~135 million years ago provides a rare opportunity to decipher the evolutionary consequences of a duplicated invertebrate genome. Here, we present a high-quality genome assembly for the mangrove horseshoe crab Carcinoscorpius rotundicauda (1.7 Gb, N50 = 90.2 Mb, with 89.8% sequences anchored to 16 pseudomolecules, 2n = 32), and a resequenced genome of the tri-spine horseshoe crab Tachypleus tridentatus (1.7 Gb, N50 = 109.7 Mb). Analyses of gene families, microRNAs, and synteny show that horseshoe crabs have undergone three rounds (3R) of WGD. Comparison of C. rotundicauda and T. tridentatus genomes from populations from several geographic locations further elucidates the diverse fates of both coding and noncoding genes. Together, the present study represents a cornerstone for improving our understanding of invertebrate WGD events on the evolutionary fates of genes and microRNAs, at both the individual and population level. We also provide improved genomic resources for horseshoe crabs, of applied value for breeding programs and conservation of this fascinating and unusual invertebrate lineage.

Post-Traumatic Brain Injury: Genetic Susceptibility to Outcome.

It is estimated that 2% of the population from industrialized countries live with lifelong disabilities resulting from traumatic brain injury (TBI) and roughly one in four adults are unable to return to work 1 year after injury because of physical or mental disabilities. TBI is a significant public health issue that causes substantial physical and economical repercussions for the individual and society. Electronic databases (PubMed, Web of Science, Google Scholar) were searched with the keywords traumatic brain injury, TBI, genes and TBI, TBI outcome, head injury. Human studies on non-penetrating traumatic brain injuries reported in English were included. To provide health care workers with the basic information for clinical management we summarize and compare the data on post-TBI outcome with regard to the impact of genetic variation: apolipoprotein E (APOE), brain-derived neurotrophic factor (BDNF), calcium channel, voltage dependent P/Q type, catechol-O-methyltransferase (COMT), dopamine receptor D2 and ankyrin repeat and kinase domain containing 1 (DRD2 and ANKK1), interleukin-1 (IL-1), interleukin-6 (IL-6), kidney and brain expressed protein (KIBRA), neurofilament, heavy polypeptide (NEFH), endothelial nitric oxide synthase 3 (NOS3), poly (ADP-ribose) polymerase-1 (PARP-1), protein phosphatase 3, catalytic subunit, gamma isozyme (PPP3CC), the serotonin transporter (5-HTT) gene solute carrier family 6 member (SLC6A4) and tumor protein 53 (TP53). It is evident that contradicting results are attributable to the heterogeneity of studies, thus further researches are warranted to effectively assess a relation between genetic traits and clinical outcome following traumatic injuries.

Evidence for differential biosynthesis of juvenile hormone (and related) sesquiterpenoids in Drosophila melanogaster.

Previous studies in Drosophila melanogaster have demonstrated that biosynthesis and regulation of juvenile hormone bisepoxide (JHB(3)) may not be coordinated with that of juvenile hormone (JH III). In this study, we have used the radiochemical assay to confirm the coordinated developmental sesquiterpenoid profile during adult life and analyze the effect of farnesol and farnesoic acid addition on methyl farnesoate, JH III and JHB(3) production by isolated ring glands of Drosophila third instar larvae or corpora allata of adult females. Application of exogenous farnesol or farnesoic acid to glands in vitro stimulated MF and JH III biosynthesis in both larvae and adults. Farnesol and farnesoic acid were inhibitory to JHB(3) biosynthesis in larvae. N-acetyl-geranyl-L-cysteine (NAGC) and S-farnesyl-thioacetic acid (SFTA) are farnesyl pyrophosphatase inhibitors that have specificity towards two different ring gland phosphatases. NAGC and SFTA had no effect on MF or JH III biosynthesis, whereas SFTA inhibited JHB(3) biosynthesis. SFTA shows specificity for a ring gland phosphatase, Phos2680, which has not been previously implicated as a contributor to JHB(3) biosynthesis. This finding suggests that farnesol production occurs in two alternate pools; one pool utilized for MF and JH III production and the other for JHB(3) production. Finally, we have used the UAS-GAL4 system in Drosophila to express juvenile hormone acid methyltransferase (JHAMT) in vivo. In contrast to in vitro studies, JHAMT expression had no effect on MF or JH III biosynthesis but stimulated JHB(3) in both larvae and adults.

Influence of codon usage bias on FGLamide-allatostatin mRNA secondary structure.

The FGLamide allatostatins (ASTs) are invertebrate neuropeptides which inhibit juvenile hormone biosynthesis in Dictyoptera and related orders. They also show myomodulatory activity. FGLamide AST nucleotide frequencies and codon bias were investigated with respect to possible effects on mRNA secondary structure. 367 putative FGLamide ASTs and their potential endoproteolytic cleavage sites were identified from 40 species of crustaceans, chelicerates and insects. Among these, 55% comprised only 11 amino acids. An FGLamide AST consensus was identified to be (X)(1→16)Y(S/A/N/G)FGLGKR, with a strong bias for the codons UUU encoding for Phe and AAA for Lys, which can form strong Watson-Crick pairing in all peptides analyzed. The physical distance between these codons favor a loop structure from Ser/Ala-Phe to Lys-Arg. Other loop and hairpin loops were also inferred from the codon frequencies in the N-terminal motif, and the first amino acids from the C-terminal motif, or the dibasic potential endoproteolytic cleavage site. Our results indicate that nucleotide frequencies and codon usage bias in FGLamide ASTs tend to favor mRNA folds in the codon sequence in the C-terminal active peptide core and at the dibasic potential endoproteolytic cleavage site.

Quantification of allatostatin receptor mRNA levels in the cockroach, Diploptera punctata, using real-time PCR.

The cockroach allatostatin receptor (Dippu-AstR) is a 425 amino acid G-protein coupled receptor that is related to the mammalian galanin receptor. Using relative standard curve real-time PCR analysis, changes in Dippu-AstR mRNA expression levels were examined in tissues of adult mated and virgin female Diploptera punctata. Tissues were chosen that were either known targets of allatostatin (Dippu-AST) action or sites of Dippu-AST localization. Tissues examined included brain, corpora allata (CA), gut, ovaries, testes and abdominal ganglia. Dippu-AstR was expressed in all tissues examined for 7 days after adult emergence. Juvenile hormone (JH) biosynthesis is known to peak on day 5 post-emergence in mated females. In mated females, Dippu-AstR mRNA was at the highest levels on day 6 post-emergence in brain and CA and day 2 post-emergence in midgut. Dippu-AstR expression was found to correlate with the decline in JH biosynthesis noted on day 5 post-emergence and early inhibition of feeding. Dippu-AstR mRNA expression in virgin female midgut and CA was dramatically elevated on days 6 and 7, respectively. Expression of Dippu-AstR mRNA was found to be similar in the abdominal ganglia of mated or virgin females. Ovarian Dippu-AstR expression declined to low levels by day 4. Testes exhibited maximal Dippu-AstR mRNA expression on days 4 and 7 of adult life. A role for Dippu-AST in testes of Diploptera is unknown.

Molecular cloning and characterization of an allatostatin-like receptor in the cockroach Diploptera punctata.

Two Drosophila receptors (AlstR/DAR-1 and DAR-2) with sequence similarity to mammalian galanin receptors have been previously identified. These receptors have been shown to form specific interactions with neuropeptides that resemble cockroach allatostatins (ASTs), which have a characteristic Tyr/Phe-Xaa-Phe-Gly-Leu-NH2 carboxyl-terminus. We hypothesized that similar allatostatin receptors exist in the cockroach Diploptera punctata that may regulate the numerous effects that this family of peptides exerts on a range of target tissues. The polymerase chain reaction (PCR) was used, with primer design based on the Drosophila allatostatin receptor (AlstR). Using these primers, a putative allatostatin-like receptor cDNA was isolated from a lambda ZAP-cDNA library prepared from the corpora allata of the D. punctata. As an approach to testing the function of this receptor in vivo, the technique of double-stranded RNA (dsRNA) gene interference was tested. Initial experiments suggest that the putative inhibition of receptor RNA expression may increase juvenile hormone (JH) production.

Quantification and visualization of Dippu-AST mRNA in the brain of adult Diploptera punctata: mated females vs. virgin females vs. males.

YXFGLa-allatostatins (ASTs) are a family of neuropeptides that inhibit juvenile hormone biosynthesis in vitro by the corpora allata of cockroaches and crickets. We have investigated the levels of AST expression in the brain of adult mated and virgin females, and adult males of the cockroach Diploptera punctata, for 11 days following emergence. To quantify Dippu-AST mRNA we have used an optimized, sensitive and specific quantitative competitive reverse-transcriptase polymerase chain reaction (QC-RT-PCR) method. Using this technique, we show that Dippu-AST is expressed at higher levels in the brains of mated females than in the brains of virgin females. Specifically, levels of Dippu-AST mRNA in day 0 mated females are 2.5x higher than levels in day 0 virgin females. By day 6-7, levels in mated females are 20x higher than those in virgin females. However, the general pattern of expression is preserved in mated and virgin females; levels of Dippu-AST mRNA are relatively high in newly eclosed females, fall steadily to significantly lower levels on day 5 (mated) and day 5-7 (virgins), and then increase and remain relatively stable until at least day 10. No significant changes in Dippu-AST mRNA are apparent in male brains during the observation period. However, the general pattern of expression is virtually the inverse of that seen in females. Dippu-AST mRNA in male brains is relatively low following emergence, increases to maximum levels on day 5, following which, levels steadily fall. In situ hybridization suggests that fluctuations in Dippu-AST mRNA expression are a result of differential transcription within medial and lateral cells of the pars intercerebralis in both males and females. In addition to yielding insights into Dippu-AST function, the quantification of Dippu-AST expression under normal conditions is useful in providing a basis for comparison of Dippu-AST expression following various physiological manipulations.

Drosophila melanogaster flatline encodes a myotropin orthologue to Manduca sexta allatostatin.

We identified a Drosophila melanogaster gene encoding a peptide that dramatically decreases spontaneous muscle contractions and, correspondingly, named the peptide flatline (FLT). This gene consisted of 4 exons and was cytologically localized to 32D2-3. Processing of a predicted 122 amino acid precursor would release pEVRYRQCYFNPISCF that differs from Manduca sexta allatostatin (Mas-AST) by one amino acid, Y4-->F4. FLT does not act as an allatostatin. In situ tissue hybridization further suggests FLT is a novel brain-gut peptide and specifically, the measured activity indicates that it is a potent myotropin. Despite its profound myotropic effect, pupae injected with FLT eclosed.