WD40 repeat proteins striatin and S/G(2) nuclear autoantigen are members of a novel family of calmodulin-binding proteins that associate with protein phosphatase 2A

Protein phosphatase 2A (PP2A) is a multifunctional serine/threonine phosphatase that is critical to many cellular processes including development, neuronal signaling, cell cycle regulation, and viral transformation. PP2A has been implicated in Ca(2+)-dependent signaling pathways, but how PP2A is targeted to these pathways is not understood. We have identified two calmodulin (CaM)-binding proteins that form stable complexes with the PP2A A/C heterodimer and may represent a novel family of PP2A B-type subunits. These two proteins, striatin and S/G(2) nuclear autoantigen (SG2NA), are highly related WD40 repeat proteins of previously unknown function and distinct subcellular localizations. Striatin has been reported to associate with the post-synaptic densities of neurons, whereas SG2NA has been reported to be a nuclear protein expressed primarily during the S and G(2) phases of the cell cycle. We show that SG2NA, like striatin, binds to CaM in a Ca(2+)-dependent manner. In addition to CaM and PP2A, several unidentified proteins stably associate with the striatin-PP2A and SG2NA-PP2A complexes. Thus, one mechanism of targeting and organizing PP2A with components of Ca(2+)-dependent signaling pathways may be through the molecular scaffolding proteins striatin and SG2NA.     

Mitochondrial DNA control region diversity in the endangered blue chaffinch, Fringilla teydea

The blue chaffinch (Fringilla teydea) is found only on the two central Canary Islands of Gran Canaria and Tenerife, where it is restricted to pine forest habitat. It is reasonably abundant on the latter island but endangered on the former. Here, sequence variation was studied in a fragment spanning domains I and II of the mitochondrial DNA (mtDNA) control region. Phylogenetic analysis of all haplotypes with a F. coelebs outgroup indicated the two island populations were reciprocally monophyletic, supporting their individual conservation. Unlike in other species, most within-island haplotype diversity was due to mutations in the domain II region. Surprisingly, genetic diversity was greater in the smaller Gran Canarian population. We suggest that this is unlikely to be maintained under current population sizes although it may be mitigated by incorporating genetic information into the captive breeding programme.     

Antiapoptotic Factor Humanin Is Expressed in Normal and Tumoral Pituitary Cells and Protects Them from TNF-α-Induced Apoptosis

Humanin (HN) is a 24-amino acid peptide with cytoprotective action in several cell types such as neurons and testicular germ cells. Rattin (HNr), a homologous peptide of HN expressed in several adult rat tissues, also has antiapoptotic action. In the present work, we demonstrated by immunocytochemical analysis and flow cytometry the expression of HNr in the anterior pituitary of female and male adult rats as well as in pituitary tumor GH3 cells. HNr was localized in lactotropes and somatotropes. The expression of HNr was lower in females than in males, and was inhibited by estrogens in pituitary cells from both ovariectomized female and orquidectomized male rats. However, the expression of HNr in pituitary tumor cells was not regulated by estrogens. We also evaluated HN action on the proapoptotic effect of TNF-α in anterior pituitary cells assessed by the TUNEL method. HN (5 µM) per se did not modify basal apoptosis of anterior pituitary cells but completely blocked the proapoptotic effect of TNF-α in total anterior pituitary cells, lactotropes and somatotropes from both female and male rats. Also, HN inhibited the apoptotic effect of TNF-α on pituitary tumor cells. In summary, our results demonstrate that HNr is present in the anterior pituitary gland, its expression showing sexual dimorphism, which suggests that gonadal steroids may be involved in the regulation of HNr expression in this gland. Antiapoptotic action of HN in anterior pituitary cells suggests that this peptide could be involved in the homeostasis of this gland. HNr is present and functional in GH3 cells, but it lacks regulation by estrogens, suggesting that HN could participate in the pathogenesis of pituitary tumors.     

Citrus tristeza virus co-opts glyceraldehyde 3-phosphate dehydrogenase for its infectious cycle by interacting with the viral-encoded protein p23

Key message

Citrus tristeza virus encodes a unique protein, p23, with multiple functional roles that include co-option of the cytoplasmic glyceraldehyde 3-phosphate dehydrogenase to facilitate the viral infectious cycle.

Abstract

The genome of citrus tristeza virus (CTV), genus Closterovirus family Closteroviridae, is a single-stranded (+) RNA potentially encoding at least 17 proteins. One (p23), an RNA-binding protein of 209 amino acids with a putative Zn-finger and some basic motifs, displays singular features: (i) it has no homologues in other closteroviruses, (ii) it accumulates mainly in the nucleolus and Cajal bodies, and in plasmodesmata, and (iii) it mediates asymmetric accumulation of CTV RNA strands, intracellular suppression of RNA silencing, induction of some CTV syndromes and enhancement of systemic infection when expressed as a transgene ectopically or in phloem-associated cells in several Citrus spp. Here, a yeast two-hybrid screening of an expression library of Nicotiana benthamiana (a symptomatic experimental host for CTV), identified a transducin/WD40 domain protein and the cytosolic glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as potential host interactors with p23. Bimolecular fluorescence complementation corroborated the p23-GAPDH interaction in planta and showed that p23 interacts with itself in the nucleolus, Cajal bodies and plasmodesmata, and with GAPDH in the cytoplasm (forming aggregates) and in plasmodesmata. The latter interaction was preserved in a p23 deletion mutant affecting the C-terminal domain, but not in two others affecting the Zn-finger and one internal basic motif. Virus-induced gene silencing of GAPDH mRNA resulted in a decrease of CTV titer as revealed by real-time RT-quantitative PCR and RNA gel-blot hybridization. Thus, like other viruses, CTV seems to co-opt GAPDH, via interaction with p23, to facilitate its infectious cycle.

     

CDK inhibitors R-roscovitine and S-CR8 effectively block renal and hepatic cystogenesis in an orthologous model of ADPKD

Autosomal dominant polycystic kidney disease (ADPKD) and other forms of PKD are associated with dysregulated cell cycle and proliferation. Although no effective therapy for the treatment of PKD is currently available, possible mechanism-based approaches are beginning to emerge. A therapeutic intervention targeting aberrant cilia-cell cycle connection using CDK-inhibitor R-roscovitine showed effective arrest of PKD in jck and cpk models that are not orthologous to human ADPKD. To evaluate whether CDK inhibition approach will translate into efficacy in an orthologous model of ADPKD, we tested R-roscovitine and its derivative S-CR8 in a model with a conditionally inactivated Pkd1 gene (Pkd1 cKO). Similar to ADPKD, Pkd1 cKO mice developed renal and hepatic cysts. Treatment of Pkd1 cKO mice with R-roscovitine and its more potent and selective analog S-CR8 significantly reduced renal and hepatic cystogenesis and attenuated kidney function decline. Mechanism of action studies demonstrated effective blockade of cell cycle and proliferation and reduction of apoptosis. Together, these data validate CDK inhibition as a novel and effective approach for the treatment of ADPKD.     

Sequence and diversity of MHC <Emphasis Type="Italic">DQA</Emphasis> and<Emphasis Type="Italic"> DQB</Emphasis> genes of the owl monkey<Emphasis Type="Italic"> Aotus nancymaae</Emphasis>

The New World primate Aotus nancymaae has been recommended by the World Health Organization (WHO) as a model for evaluation of malaria vaccine candidates, given its susceptibility to experimental infection with the human malaria parasites Plasmodium falciparum and Plasmodium vivax. We present here the nucleotide sequences of the complete cDNA of MHC-DQA1 and of the polymorphic exon 2 segments of MHC-DQB1/DQB2. In a group of three nonrelated animals captured in the wild, five alleles of MHC-DQA1 could be identified. They all belong to one lineage, namely Aona-DQA1*27. This lineage has not been described in any other New World monkey species studied. In a group of 19 unrelated animals, 14 Aona-DQB1 alleles could be identified which are grouped into the two lineages Aona-DQB1*22 and Aona-DQB1*23. These lineages have been described previously in the common marmoset and cotton-top tamarin. In addition, two Aona-DQB2 sequences could be identified which are highly similar to HLA-DQB2 sequences. Essential amino acid residues contributing to MHC DQ peptide binding pockets number 1 and 4 are conserved or semi-conserved between HLA-DQ and Aona-DQ molecules, indicating a capacity to bind similar peptide repertoires. These results fully support the use of Aotus monkeys as an animal model for evaluation of future subunit vaccine candidates.     

Distinct population structure for co-occurring Anopheles goeldii and Anopheles triannulatus in Amazonian Brazil

To evaluate whether environmental heterogeneity contributes to the genetic heterogeneity in Anopheles triannulatus, larval habitat characteristics across the Brazilian states of Roraima and Pará and genetic sequences were examined. A comparison with Anopheles goeldii was utilised to determine whether high genetic diversity was unique to An. triannulatus. Student t test and analysis of variance found no differences in habitat characteristics between the species. Analysis of population structure of An. triannulatus and An. goeldii revealed distinct demographic histories in a largely overlapping geographic range. Cytochrome oxidase I sequence parsimony networks found geographic clustering for both species; however nuclear marker networks depicted An. triannulatus with a more complex history of fragmentation, secondary contact and recent divergence. Evidence of Pleistocene expansions suggests both species are more likely to be genetically structured by geographic and ecological barriers than demography. We hypothesise that niche partitioning is a driving force for diversity, particularly in An. triannulatus.