Trio Haploinsufficiency Causes Neurodevelopmental Disease-Associated Deficits

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Mass spectral analyses of hemoglobin adducts formed after in vitro exposure of erythrocytes to hydroxymethylvinyl ketone

Hydroxymethylvinyl ketone (HMVK) is a reactive oxidation product of 3-butene-1,2-diol, a metabolite of 1,3-butadiene. The potential for HMVK (0.1 and 1mM) to form hemoglobin (Hb) adducts in erythrocytes from Sprague-Dawley rats was investigated at physiological conditions (pH 7.4, 37 degrees C) using electrospray ionization mass spectrometry (ESI/MS). With the 0.1mM HMVK globin samples, the results indicate HMVK adduction on the alpha2, beta2 and beta3 chains. With the 1.0mM HMVK globin samples, adducts were detected on the beta2 and beta3 chains. However, no correlation was observed between incubation time and the extent of adduct formation, and additional adducts were detected when globin samples were fractionated by HPLC before the ESI/MS analyses. For specific localizations of adducts on the globin chains, trypsin digested peptides from the 1mM HMVK globin samples were subjected to liquid chromatography/mass spectrometry analyses. The results, which are consistent with formation of HMVK adducts on several specific peptides within the alpha- and beta-chains, suggest selectivity in the interaction of HMVK with the different cysteine residues in Hb. Because adducts were also detected in peptides containing no cysteine residues and multiple HMVK moieties were detected on some of the cysteine-containing peptides, the results suggest other amino acids may be also reactive with HMVK. Adduct profiles and their relative intensities were consistent between the 1 and 2h samples providing evidence for the HMVK reactions being fast and selective. The finding that fewer peptides were adducted in the 0.1mM HMVK globin samples provides further evidence for selectivity of the HMVK reaction. Collectively, the results show HMVK readily and selectively forms adducts on Hb. Characterization of these adducts will facilitate development of useful biomarkers of exposure to HMVK and its precursor 1,3-butadiene.     

ProSight Annotator: Complete control and customization of protein entries in UniProt XML files

The effectiveness of any proteomics database search depends on the theoretical candidate information contained in the protein database. Unfortunately, candidate entries from protein databases such as UniProt rarely contain all the post-translational modifications (PTMs), disulfide bonds, or endogenous cleavages of interest to researchers. These omissions can limit discovery of novel and biologically important proteoforms. Conversely, searching for a specific proteoform becomes a computationally difficult task for heavily modified proteins. Both situations require updates to the database through user-annotated entries. Unfortunately, manually creating properly formatted UniProt Extensible Markup Language (XML) files is tedious and prone to errors. ProSight Annotator solves these issues by providing a graphical interface for adding user-defined features to UniProt-formatted XML files for better informed proteoform searches. It can be downloaded from http://prosightannotator.northwestern.edu .     

Unique sympatric quartet of limnetic,benthic, profundal and piscivorous brown trout populations resolved by 3D sampling and focused molecular marker selection

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Alterations in habituation of the tail flip response in epigean and troglobitic crayfish

We demonstrate that the probability of the crayfish, P. clarkii, to tail flip in response to a touch on the dorsal tail fan is dependent on both the size and the behavioral state of the animal. Alterations in the animal's internal physical state, such as when the animal autotomizes its chelipeds, will cause larger-sized animals to tail flip; if they were not autotomized, then no tail flip response would occur. Altering the external environment by removal of water causes small crayfish, which normally habituate slowly, to rapidly habituate. Observation of large adult crayfish in a species, O. australis packardi, one that evolved to live in total cave darkness, revealed that they are more likely to tail flip than are the sighted, adult P. clarkii. Results indicate that the behavioral state of the crayfish can result in rapid and long-term alterations in the tail flip response and in habituation rates to repetitive stimuli. This ability to show plasticity in gain setting may be regulated by neuromodulators and can occur in large adults of the sighted crayfish. Differences between the two species indicate that size may not be the sole contributing factor to account for tail flip behaviors. J. Exp. Zool. 290:163-176, 2001.     

Thiopalmitoylation of myelin proteolipid protein epitopes enhances immunogenicity and encephalitogenicity

Proteolipid protein (PLP) is the most abundant protein of CNS myelin, and is posttranslationally acylated by covalent attachment of long chain fatty acids to cysteine residues via a thioester linkage. Two of the acylation sites are within epitopes of PLP that are encephalitogenic in SJL/J mice (PLP(104-117) and PLP(139-151)) and against which increased immune responses have been detected in some multiple sclerosis patients. It is known that attachment of certain types of lipid side chains to peptides can result in their enhanced immunogenicity. The aim of this study was to determine whether thioacylated PLP peptides, as occur in the native protein, are more immunogenic than their nonacylated counterparts, and whether thioacylation influences the development of autoreactivity and experimental autoimmune encephalomyelitis. The results show that in comparison with nonacylated peptides, thioacylated PLP lipopeptides can induce greater T cell and Ab responses to both the acylated and nonacylated peptides. They also enhanced the development and chronicity of experimental autoimmune encephalomyelitis. Synthetic peptides in which the fatty acid was attached via an amide linkage at the N terminus were not encephalitogenic, and they induced greater proportions of CD8+ cells in initial in vitro stimulation. Therefore, the lability and the site of the linkage between the peptide and fatty acid may be important for induction of encephalitogenic CD4+ T cells. These results suggest that immune responses induced by endogenous thioacylated lipopeptides may contribute to the immunopathogenesis of chronic experimental demyelinating diseases and multiple sclerosis.     

Identification of a novel gene, DZIP (DAZ-interacting protein), that encodes a protein that interacts with DAZ (deleted in azoospermia) and is expressed in embryonic stem cells and germ cells

Evidence from diverse organisms, including humans, suggests that the DAZ (Deleted in Azoospermia) gene and a closely related homolog, DAZL (DAZ-like), are required early in germ cell development to maintain initial germ cell populations. Here we report the identification and characterization of the DZIP (DAZ-Interacting Protein) gene, which encodes at least three different protein isoforms that contain a C2H2 zinc-finger domain. The DZIP gene is expressed predominantly in human embryonic stem cells and fetal and adult germ cells; moreover, two DZIP protein isoforms colocalize with DAZ and/or DAZL proteins in these tissues. Finally, we provide evidence indicating that DZIP may associate with DAZ and its other cofactors in an RNA-binding protein complex that functions in both ES cells and germ cells.     

Interannual Variation in Timing of Parturition and Growth of Collared Pikas (Ochotona collaris) in the Southwest Yukon

The length of the snow-free season has a significant influenceon reproduction and growth in northern alpine environments,and these life history traits may provide sensitive indicatorsof the responses of organisms to climate change. We examinedgrowth rates and timing of parturition of collared pikas (Ochotonacollaris) from 1995–2002 in the Ruby Range, Yukon Territory,Canada. Growth rates were best described using a Gompertz model,in which the asymptotic mass, determined from the average maleand female weights, was 157 g, the growth rate constant (K)was 0.0557, and the age at inflection (I) was 18.12 days, fora birth weight of 10 g. The maximum growth rate for North Americanpikas (O. collaris and O. princeps) increased with latitude,with maximum growth rates being approximately one-third greaterin northern populations where the snow-free season is less thanthree months long. The mean parturition date varied significantlyamong years from 3 June to 3 July, and delayed parturition wascorrelated with indices of high snow accumulation and, to alesser extent, late spring snowmelt. However, parturition datedid not significantly affect the subsequent over-winter survivalof juveniles in this population, suggesting that pikas are ableto adjust to seasonal uncertainty associated with highly variablespring conditions.