The impact of coral bleaching on the pigment profile of the symbiotic alga, Symbiodinium

Bleaching of corals by loss of symbiotic dinoflagellate algae and/or photosynthetic pigments is commonly triggered by elevated temperatures coupled with high irradiance, and is a first-order threat to coral reef communities. In this study, a high-resolution high-performance liquid chromatography method integrated with mass spectrometry was applied to obtain the first definitive identification of chlorophyll and carotenoid pigments of three clades of symbiotic dinoflagellate algae (Symbiodinium) in corals, and their response to experimentally elevated temperature and irradiance. The carotenoids peridinin, dinoxanthin, diadinoxanthin (Dn), diatoxanthin (Dt) and beta-carotene were detected, together with chlorophylls a and c2, and phaeophytin a, in all three algal clades in unstressed corals. On exposure to elevated temperature and irradiance, three coral species (Montastrea franksi and Favia fragum with clade B algae, and Montastrea cavernosa with clade C) bleached by loss of 50-80% of their algal cells, with no significant impact to chlorophyll a or c2, or peridinin in retained algal cells. One species (Agaricia sp. with clade C) showed no significant reduction in algal cells at elevated temperature and irradiance, but lost substantial amounts of chlorophyll a and carotenoid pigments, presumably through photo-oxidative processes. Two coral species (Porites astreoides and Porites porites both bearing clade A algae) did not bleach. The impact of elevated temperature and irradiance on the levels of the photoprotective xanthophylls (Dn + Dt) and beta-carotene varied among the corals, both in pool size and xanthophyll cycling, and was not correlated to coral bleaching resistance.     

S6K1 regulates GSK3 under conditions of mTOR-dependent feedback inhibition of Akt

Feedback inhibition of the PI3K-Akt pathway by the mammalian target of rapamycin complex 1 (mTORC1) has emerged as an important signaling event in tumor syndromes, cancer, and insulin resistance. Cells lacking the tuberous sclerosis complex (TSC) gene products are a model for this feedback regulation. We find that, despite Akt attenuation, the Akt substrate GSK3 is constitutively phosphorylated in cells and tumors lacking TSC1 or TSC2. In these settings, GSK3 phosphorylation is sensitive to mTORC1 inhibition by rapamycin or amino acid withdrawal, and GSK3 becomes a direct target of S6K1. This aberrant phosphorylation leads to decreased GSK3 activity and phosphorylation of downstream substrates and contributes to the growth-factor-independent proliferation of TSC-deficient cells. We find that GSK3 can also be regulated downstream of mTORC1 in a HepG2 model of cellular insulin resistance. Therefore, we define conditions in which S6K1, rather than Akt, is the predominant GSK3 regulatory kinase.     

Functional consequence of positive selection revealed through rational mutagenesis of human myeloperoxidase

Myeloperoxidase (MPO) is a member of the mammalian heme peroxidase (MHP) multigene family. Whereas all MHPs oxidize specific halides to generate the corresponding hypohalous acid, MPO is unique in its capacity to oxidize chloride at physiologic pH to produce hypochlorous acid (HOCl), a potent microbicide that contributes to neutrophil-mediated host defense against infection. We have previously resolved the evolutionary relationships in this functionally diverse multigene family and predicted in silico that positive Darwinian selection played a major role in the observed functional diversities (Loughran NB, O'Connor B, O'Fagain C, O'Connell MJ. 2008. The phylogeny of the mammalian heme peroxidases and the evolution of their diverse functions. BMC Evol Biol. 8:101). In this work, we have replaced positively selected residues asparagine 496 (N496), tyrosine 500 (Y500), and leucine 504 (L504) with the amino acids present in the ancestral MHP and have examined the effects on the structure, biosynthesis, and activity of MPO. Analysis in silico predicted that N496F, Y500F, or L504T would perturb hydrogen bonding in the heme pocket of MPO and thus disrupt the structural integrity of the enzyme. Biosynthesis of the mutants stably expressed in human embryonic kidney 293 cells yielded apoproMPO, the heme-free, enzymatically inactive precursor of MPO, that failed to undergo normal maturation or proteolytic processing. As a consequence of the maturational arrest at the apoproMPO stage of development, cells expressing MPO with mutations N496F, Y500F, L504T, individually or in combination, lacked normal peroxidase or chlorinating activity. Taken together, our data provide further support for the in silico predictions of positive selection and highlight the correlation between positive selection and functional divergence. Our data demonstrate that directly probing the functional importance of positive selection can provide important insights into understanding protein evolution.     

Population-based modeling of the progression of apoptosis in mammalian cell culture

The production of biopharmaceuticals from mammalian cell culture is hindered by apoptosis, which is the primary cause of cell death in these cultures. As a tool for optimization of culture yield, this study presents a population-based model describing the progression of apoptosis in a monoclonal antibody (mAb)-producing Chinese hamster ovary (CHO) cell culture. Because mAb production does not cease when apoptosis begins, the model was designed to incorporate subpopulations at various stages in the progression of apoptosis. The model was validated against intracellular measurements of caspase activity as well as cell density, nutrient levels, and toxic metabolites. Since the specific details of apoptotic mechanisms have not been elucidated in this cell line, we employed a model comparison analysis that suggests the most plausible pathways of activation.     

The histone variant macroH2A1.1 is recruited to DSBs through a mechanism involving PARP1

The repair of DNA double-strand breaks (DSBs) requires remodeling of the local chromatin architecture to allow the repair machinery to access sites of damage. Here, we report that the histone variant macroH2A1.1 is recruited to DSBs. Cells lacking macroH2A1 have defective recruitment of 53BP1, defective activation of chk2 kinase and increased radiosensitivity. Importantly, macroH2A1.1 is not incorporated into nucleosomes at DSBs, but instead associates with the chromatin through a mechanism which requires PARP1 activity. These results reveal an unusual mechanism involving a direct association of macroH2A1.1 with PARylated chromatin which is critical for retaining 53BP1 at sites of damage.     

Ultraconserved elements in the human genome: association and transmission analyses of highly constrained single-nucleotide polymorphisms

Ultraconserved elements in the human genome likely harbor important biological functions as they are dosage sensitive and are able to direct tissue-specific expression. Because they are under purifying selection, variants in these elements may have a lower frequency in the population but a higher likelihood of association with complex traits. We tested a set of highly constrained SNPs (hcSNPs) distributed genome-wide among ultraconserved and nearly ultraconserved elements for association with seven traits related to reproductive (age at natural menopause, number of children, age at first child, and age at last child) and overall [longevity, body mass index (BMI), and height] fitness. Using up to 24,047 European-American samples from the National Heart, Lung, and Blood Institute Candidate Gene Association Resource (CARe), we observed an excess of associations with BMI and height. In an independent replication panel the most strongly associated SNPs showed an 8.4-fold enrichment of associations at the nominal level, including three variants in previously identified loci and one in a locus (DENND1A) previously shown to be associated with polycystic ovary syndrome. Finally, using 1430 family trios, we showed that the transmissions from heterozygous parents to offspring of the derived alleles of rare (frequency ≤0.5%) hcSNPs are not biased, particularly after adjusting for the rates of genotype missingness and error in the data. The lack of transmission bias ruled out an immediately and strongly deleterious effect due to the rare derived alleles, consistent with the observation that mice homozygous for the deletion of ultraconserved elements showed no overt phenotype. Our study also illustrated the importance of carefully modeling potential technical confounders when analyzing genotype data of rare variants.     

Genome sequence of the halophilic archaeon Halococcus hamelinensis

Halococcus hamelinensis was isolated from hypersaline stromatolites in Shark Bay, Australia. Here we report the genome sequence (3,133,046 bp) of H. hamelinensis, which provides insights into the ecology, evolution, and adaptation of this novel microorganism.