The effect of extreme weather events on breeding parameters of the White Stork Ciconia ciconia

Capsule: Although the White Stork avoids adverse weather conditions by modifying its arrival and breeding, it cannot avoid extreme weather events during the breeding season. Aims: To show how extreme weather conditions can influence breeding attempts of a large, long-lived species, the White Stork. Methods: We analysed data on arrivals of White Storks in Western Poland from 2005 to 2013 and detailed breeding biology parameters from 2009 to 2013 in relation to weather conditions. We analysed breeding success and breeding failure rate from 1974 to 2013. Results: In years with a cold March White Storks arrived later than when March was warmer. Frost during incubation negatively influenced the hatching success. Extreme weather events caused high late mortality even for nestlings older than 30 days. Data from 27 breeding seasons showed a significant increase in mean breeding success but also a significant increase in the proportion of pairs which lost broods in the nestling stage. Conclusion: The White Stork can modify its arrival in response to current weather conditions on the breeding grounds but it cannot respond to extreme weather events. Due to increasing frequency of extreme weather events caused by climate change, White Stork breeding success may decrease in the future. Publisher statement: This is an Accepted Manuscript of an article published by Taylor & Francis in Bird Study on 24th July 2015, available online: http://www.tandfonline.com/10.1080/00063657.2015.105874

Protein phosphatase beta, a putative type-2A protein phosphatase from the human malaria parasite Plasmodium falciparum.

Protein phosphatases play a critical role in the regulation of the eukaryotic cell cycle and signal transduction. A putative protein serine/threonine phosphatase gene has been isolated from the human malaria parasite Plasmodium falciparum. The gene has an unusual intron that contains four repeats of 32 nucleotides and displays a high degree of size polymorphism among different strains of P. falciparum. The open reading frame reconstituted by removal of the intron encodes a protein of 466 amino acids with a predicted molecular mass of approximately 53.7 kDa. The encoded protein, termed protein phosphatase beta (PP-beta), is composed of two distinct domains. The C-terminal domain comprises 315 amino acids and exhibits a striking similarity to the catalytic subunits of the type-2A protein phosphatases. Database searches revealed that the catalytic domain has the highest similarity to Schizosaccharomyces pombe Ppa1 (58% identity and 73% similarity). However, it contains a hydrophilic insert consisting of five amino acids. The N-terminal domain comprises 151 amino acid residues and exhibits several striking features, including high levels of charged amino acids and asparagine, and multiple consensus phosphorylation sites for a number of protein kinases. An overall structural comparison of PP-beta with other members of the protein phosphatase 2A group revealed that PP-beta is more closely related to Saccharomyces cerevisiae PPH22. Southern blots of genomic DNA digests and chromosomal separations showed that PP-beta is a single-copy gene and is located on chromosome 9. A 2800-nucleotide transcript of this gene is expressed specifically in the sexual erythrocytic stage (gametocytes). The results indicate that PP-beta may be involved in sexual stage development

Monitoring cell proliferation in vitro with different cellular fluorescent dyes

 There are few methods for quantifying cell proliferation. Those tests describe the proliferation kinetics of a cell population, but they do not report the history of single cells, the number and frequency of cell divisions, or the precursor cell frequency. Cell-tracking assays based on dilution of the green fluorescent protein labelling dye, CFSE, has become the standard for monitoring cell proliferation. Other labelling dyes, e.g. CellTrace Violet and CellVue Claret, are also used for the same purpose. This study aimed to compare these three cell labelling methods for analysing the kinetics of cell viability, proliferation, and precursor cell frequency. Human peripheral blood mononuclear cells stimulated with Concanavalin A (ConA) were used as a model system. After labelling with a cell-tracking dye cells were divided into groups with and without ConA stimulation. From the 5th to 8th day, cells were collected and analysed with flow cytometry. Cell viability was not significantly different between labelled and unlabelled cells that received ConA stimulation. The proliferative fraction, proliferation index, and nonproliferative fraction were not significantly different among lymphocytes labelled with different dyes. Precursor cell frequency was also similar among cells labelled with the three cell-tracing dyes. The practical conclusion from our observations is that the results from cells labelled with different tracers may be compared directly and discussed jointly.

Focal Cerebral Ischemia Reduces Protein Phosphatase 2A Subunit B Expression in Brain Tissue and HT22 Cells

Protein phosphatase 2A (PP2A) is a serine and threonine protein phosphatase that regulates cell cycle progression and apoptosis. PP2A is composed of various subunits. Among these subunits, subunit B plays an important role in the modulation of PP2A function in the brain. This study investigated PP2A subunit B expression levels after neuronal cell injury. Middle cerebral artery occlusions (MCAO) were surgically induced in adult male rats to induce focal cerebral ischemic injury, and brain tissues were collected 24 h after MCAO. A proteomic approach revealed reduction of PP2A subunit B protein spots in MCAO-operated animals in comparison to sham-operated animals. Western blot analysis confirmed that MCAO induces reductions in PP2A subunit B levels. Moreover, glutamate exposure induces neuronal cell death and leads to reductions of PP2A subunit B levels in a hippocampal-derived cell line. This study demonstrated the decrease of PP2A subunit B in ischemic neuronal cell injury. These results suggest that the decrease of PP2A subunit B after ischemic brain injury can mediate neuronal cell death

Characterisation of the Mouse Vasoactive Intestinal Peptide Receptor Type 2 Gene, Vipr2, and Identification of a Polymorphic LINE-1-like Sequence That Confers Altered Promoter Activity

The VPAC(2) receptor is a seven transmembrane spanning G protein-coupled receptor for two neuropeptides, vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP). It has a distinct tissue-specific, developmental and inducible expression that underlies an important neuroendocrine role. Here, we report the characterisation of the gene that encodes the mouse VPAC(2) receptor (Vipr2), localisation of the transcriptional start site and functional analysis of the promoter region. The Vipr2 gene contains 12 introns within its protein-coding region and spans 68.6 kb. Comparison of the 5′ untranslated region sequences for cloned 5′-RACE products amplified from different tissues showed they all were contained within the same exon, with the longest extending 111 bp upstream of the ATG start site. Functional analysis of the 3.2-kb 5′-flanking region using sequentially deleted sequences cloned into a luciferase gene reporter vector revealed that this region is active as a promoter in mouse AtT20 D16:16 and rat GH4C1 cell lines. The core promoter is located within a 180-bp GC-rich region proximal to the ATG start codon and contains potential binding sites for Sp1 and AP2, but no TATA-box. Further upstream, in two out of three mice strains examined, we have discovered a 496-bp polymorphic DNA sequence that bears a significant identity to mouse LINE-1 DNA. Comparison of the promoter activity between luciferase reporter gene constructs derived from the BALB/c (which contains this sequence) and C57BL/6J (which lacks this sequence) Vipr2 promoter regions has shown three-fold difference in luciferase gene activity when expressed in mouse AtT20 D16:16 and αT3-1 cells, but not when expressed in the rat GH4C1 cells or in COS 7 cells. Our results suggest that the mouse Vipr2 gene may be differentially active in different mouse strains, depending on the presence of this LINE-1-like sequence in the promoter region

The variable subunit associated with protein phosphatase 2A0 defines a novel multimember family of regulatory subunits

Two protein phosphatase 2A (PP2A) holoenzymes were isolated from rabbit skeletal muscle containing, in addition to the catalytic and PR65 regulatory subunits, proteins of apparent molecular masses of 61 and 56 kDa respectively. Both holoenzymes displayed low basal phosphorylase phosphatase activity, which could be stimulated by protamine to an extent similar to that of previously characterized PP2A holoenzymes. Protein microsequencing of tryptic peptides derived from the 61 kDa protein, termed PR61, yielded 117 residues of amino acid sequence. Molecular cloning by enrichment of specific mRNAs, followed by reverse transcription–PCR and cDNA library screening, revealed that this protein exists in multiple isoforms encoded by at least three genes, one of which gives rise to several splicing variants. Comparisons of these sequences with the available databases identified one more human gene and predicted another based on a rabbit cDNA-derived sequence, thus bringing the number of genes encoding PR61 family members to five. Peptide sequences derived from PR61 corresponded to the deduced amino acid sequences of either α or β isoforms, indicating that the purified PP2A preparation was a mixture of at least two trimers. In contrast, the 56 kDa subunit (termed PR56) seems to correspond to the ϵ isoform of PR61. Several regulatory subunits of PP2A belonging to the PR61 family contain consensus sequences for nuclear localization and might therefore target PP2A to nuclear substrates.</jats:p

Protein phosphatase 2A plays a crucial role in Giardia lamblia differentiation

Author Posting. © The Authors, 2006. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Molecular and Biochemical Parasitology 152 (2007): 80-89, doi:10.1016/j.molbiopara.2006.12.001.The ability of Giardia lamblia to undergo two distinct differentiations in response to physiologic stimuli is central to its pathogenesis. The giardial cytoskeleton changes drastically during encystation and excystation. However, the signal transduction pathways mediating these transformations are poorly understood. We tested the hypothesis that PP2A, a highly conserved serine/threonine protein phosphatase, might be important in giardial differentiation. We found that in vegetatively growing trophozoites, gPP2A-C protein localizes to basal bodies/centrosomes, and to cytoskeletal structures unique to Giardia: the ventral disk, and the dense rods of the anterior, posterior-lateral, and caudal flagella. During encystation, gPP2A-C protein disappears from only the anterior flagellar dense rods. During excystation, gPP2A-C localizes to the cyst wall in excysting cysts but is not found in the wall of cysts with emerging excyzoites. Transcriptome and immunoblot analyses indicated that gPP2A-C mRNA and protein are upregulated in mature cysts and during the early stage of excystation that models passage through the host stomach. Stable expression of gPP2A-C antisense RNA did not affect vegetative growth, but strongly inhibited the formation of encystation secretory vesicles (ESV) and water-resistant cysts. Moreover, the few cysts that formed were highly defective in excystation. Thus, gPP2A-C localizes to universal cytoskeletal structures and to structures unique to Giardia. It is also important for encystation and excystation, crucial giardial transformations that entail entry into and exit from dormancy.This work was funded by NIH grants GM61896, AI51687, AI42488, and DK35108. Dr. A.G. McArthur was supported by NIH grant AI51089 and the Marine Biological Laboratory’s Program in Global Infectious Diseases, funded by the Ellison Medical Foundation

Essential Roles of the Tap42-Regulated Protein Phosphatase 2A (PP2A) Family in Wing Imaginal Disc Development of Drosophila melanogaster

Protein ser/thr phosphatase 2A family members (PP2A, PP4, and PP6) are implicated in the control of numerous biological processes, but our understanding of the in vivo function and regulation of these enzymes is limited. In this study, we investigated the role of Tap42, a common regulatory subunit for all three PP2A family members, in the development of Drosophila melanogaster wing imaginal discs. RNAi-mediated silencing of Tap42 using the binary Gal4/UAS system and two disc drivers, pnr- and ap-Gal4, not only decreased survival rates but also hampered the development of wing discs, resulting in a remarkable thorax cleft and defective wings in adults. Silencing of Tap42 also altered multiple signaling pathways (HH, JNK and DPP) and triggered apoptosis in wing imaginal discs. The Tap42RNAi-induced defects were the direct result of loss of regulation of Drosophila PP2A family members (MTS, PP4, and PPV), as enforced expression of wild type Tap42, but not a phosphatase binding defective Tap42 mutant, rescued fly survivorship and defects. The experimental platform described herein identifies crucial roles for Tap42•phosphatase complexes in governing imaginal disc and fly development