Posttranslational modification of proteins by acetylation and phosphorylation regulates most cellular processes in living organisms. Surprisingly, the evolutionary conservation of phosphorylated serine and threonine residues is only marginally higher than that of unmodified serines and threonines. With high-resolution mass spectrometry, we identified 1981 lysine acetylation sites in the proteome of Drosophila melanogaster. We used data sets of experimentally identified acetylation and phosphorylation sites in Drosophila and humans to analyze the evolutionary conservation of these modification sites between flies and humans. Site-level conservation analysis revealed that acetylation sites are highly conserved, significantly more so than phosphorylation sites. Furthermore, comparison of lysine conservation in Drosophila and humans with that in nematodes and zebrafish revealed that acetylated lysines were significantly more conserved than were nonacetylated lysines. Bioinformatics analysis using Gene Ontology terms suggested that the proteins with conserved acetylation control cellular processes such as protein translation, protein folding, DNA packaging, and mitochondrial metabolism. We found that acetylation of ubiquitin-conjugating E2 enzymes was evolutionarily conserved, and mutation of a conserved acetylation site impaired the function of the human E2 enzyme UBE2D3. This systems-level analysis of comparative posttranslational modification showed that acetylation is an anciently conserved modification and suggests that phosphorylation sites may have evolved faster than acetylation sites.

Post-translational modification of proteins by lysine acetylation plays important regulatory roles in living cells. The budding yeast Saccharomyces cerevisiae is a widely used unicellular eukaryotic model organism in biomedical research. S. cerevisiae contains several evolutionary conserved lysine acetyltransferases and deacetylases. However, only a few dozen acetylation sites in S. cerevisiae are known, presenting a major obstacle for further understanding the regulatory roles of acetylation in this organism. Here we use high resolution mass spectrometry to identify about 4000 lysine acetylation sites in S. cerevisiae. Acetylated proteins are implicated in the regulation of diverse cytoplasmic and nuclear processes including chromatin organization, mitochondrial metabolism, and protein synthesis. Bioinformatic analysis of yeast acetylation sites shows that acetylated lysines are significantly more conserved compared with nonacetylated lysines. A large fraction of the conserved acetylation sites are present on proteins involved in cellular metabolism, protein synthesis, and protein folding. Furthermore, quantification of the Rpd3-regulated acetylation sites identified several previously known, as well as new putative substrates of this deacetylase. Rpd3 deficiency increased acetylation of the SAGA (Spt-Ada-Gcn5-Acetyltransferase) complex subunit Sgf73 on K33. This acetylation site is located within a critical regulatory domain in Sgf73 that interacts with Ubp8 and is involved in the activation of the Ubp8-containing histone H2B deubiquitylase complex. Our data provides the first global survey of acetylation in budding yeast, and suggests a wide-ranging regulatory scope of this modification. The provided dataset may serve as an important resource for the functional analysis of lysine acetylation in eukaryotes.

DNA replication stress is a source of genomic instability. Here we identify changed mutation rate 1 (Cmr1) as a factor involved in the response to DNA replication stress in Saccharomyces cerevisiae and show that Cmr1—together with Mrc1/Claspin, Pph3, the chaperonin containing TCP1 (CCT) and 25 other proteins—define a novel intranuclear quality control compartment (INQ) that sequesters misfolded, ubiquitylated and sumoylated proteins in response to genotoxic stress. The diversity of proteins that localize to INQ indicates that other biological processes such as cell cycle progression, chromatin and mitotic spindle organization may also be regulated through INQ. Similar to Cmr1, its human orthologue WDR76 responds to proteasome inhibition and DNA damage by relocalizing to nuclear foci and physically associating with CCT, suggesting an evolutionarily conserved biological function. We propose that Cmr1/WDR76 plays a role in the recovery from genotoxic stress through regulation of the turnover of sumoylated and phosphorylated proteins.

A manipulative mesocosm experiment in Danish coastal waters tested the effect on plankton biodiversity and function of adding nitrate, phosphate and glucose. A comprehensive set of measurements was made over a 6 d period; these included phytoplankton biomass and production in 3 size fractions (>10, 10-2 and < 2 µm), bacterial biomass and production, nitrate and ammonium uptake, and pigment taxonomy. Addition of nitrate and phosphate resulted in increases of biomass and production of all size fractions of phytoplankton. Inorganic nutrients alone had only a minor effect on bacterial abundance and production, with slight increases relative to the control. The largest changes occurred in mesocosms to which glucose was added in excess with nitrate and phosphate. Pigment composition indicated little change in phytoplankton assemblage composition in any treatment. A large increase in bacterial activity in the presence of added glucose had a negative effect on the phytoplankton assemblage and resulted in a decline in phytoplankton biomass. Data on nutrient uptake and size-fractionated carbon fixation suggest that the mechanism of this phytoplankton suppression was the ability of heterotrophic bacteria to out-compete for available inorganic nutrients, resulting in nutrient limitation of the phytoplankton assemblage. KEY WORDS: Phytoplankton/bacterial competition · Nutrient uptake · Pigment taxonomy · MesocosmResale or republication not permitted without written consent of the publisher

Light and electron microscopy, nuclear-encoded LSU rDNA sequences, and pigment analyses were performed on five geographically separate isolates of Gymnodinium mikimotoi. The morphological variation between the isolates equals that found within the isolates. The nuclear-encoded LSU rDNA sequences were nearly identical in all isolates, and molecular analyses using maximum likelihood, parsimony, and neighbor joining showed the geographical isolates as an unresolved clade. Based on the available data it is concluded that the European isolates, formerly identified as Gyrodinium aureolum , Gyrodinium cf. aureolum , or Gymnodinium cf. nagasakiense , are conspecific with the Japanese Gymnodinium mikimotoi. An isolate from the Pettaquamscutt River, USA, is suggested to represent what Hulburt (1957) described as Gyrodinium aureolum. The LSU rDNA sequence data and ultrastructural characters in this isolate closely resemble those of Gymnodinium fuscum , the type species of Gymnodinium , and Gyrodinium aureolum Hulburt is therefore renamed Gymnodinium aureolum (Hulburt) G. Hansen, comb. nov.

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The diversity of prokaryotic and eukaryotic phytoplankton was studied along a gradient of salinity in the solar salterns of Bras del Port in Santa Pola (Alacant, Spain) using different community descriptors. Chlorophyll a, HPLC pigment composition, flow cytometrically-determined picoplankton concentration, taxonomic composition of phytoplankton (based on optical microscopy) and genetic fingerprint patterns of 16S (cyanobacteria- and chloroplast-specific primers) and 18S rRNA genes were determined for samples from ponds with salinities ranging from 4% to 37%. Both morphological and genetical descriptors of taxonomic composition showed a good agreement and indicated a major discontinuity at salinities between 15% and 22%. The number of classes and the Shannon diversity index corresponding to the different descriptors showed a consistent decreasing trend with increasing salinity. The results indicate a selective effect of extremely high salinities on phytoplanktonic assemblages.

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