No abstract

Prokaryotes, due to their moderate complexity, are particularly amenable to the comprehensive identification of the protein repertoire expressed under different conditions. We applied a generic strategy to identify a complete expressed prokaryotic proteome, which is based on the analysis of RNA and proteins extracted from matched samples. Saturated transcriptome profiling by RNA-seq provided an endpoint estimate of the protein-coding genes expressed under two conditions which mimic the interaction of Bartonella henselae with its mammalian host. Directed shotgun proteomics experiments were carried out on four subcellular fractions. By specifically targeting proteins which are short, basic, low abundant, and membrane localized, we could eliminate their initial underrepresentation compared to the estimated endpoint. A total of 1250 proteins were identified with an estimated false discovery rate below 1%. This represents 85% of all distinct annotated proteins and 90% of the expressed protein-coding genes. Genes that were detected at the transcript but not protein level, were found to be highly enriched in several genomic islands. Furthermore, genes that lacked an ortholog and a functional annotation were not detected at the protein level; these may represent examples of overprediction in genome annotations. A dramatic membrane proteome reorganization was observed, including differential regulation of autotransporters, adhesins, and hemin binding proteins. Particularly noteworthy was the complete membrane proteome coverage, which included expression of all members of the VirB/D4 type IV secretion system, a key virulence factor.

During the Early Bronze Age, populations of the western Eurasian steppe expanded across an immense area of northern Eurasia. Combined archaeological and genetic evidence supports widespread Early Bronze Age population movements out of the Pontic–Caspian steppe that resulted in gene flow across vast distances, linking populations of Yamnaya pastoralists in Scandinavia with pastoral populations (known as the Afanasievo) far to the east in the Altai Mountains1,2 and Mongolia3. Although some models hold that this expansion was the outcome of a newly mobile pastoral economy characterized by horse traction, bulk wagon transport4–6 and regular dietary dependence on meat and milk5, hard evidence for these economic features has not been found. Here we draw on proteomic analysis of dental calculus from individuals from the western Eurasian steppe to demonstrate a major transition in dairying at the start of the Bronze Age. The rapid onset of ubiquitous dairying at a point in time when steppe populations are known to have begun dispersing offers critical insight into a key catalyst of steppe mobility. The identification of horse milk proteins also indicates horse domestication by the Early Bronze Age, which provides support for its role in steppe dispersals. Our results point to a potential epicentre for horse domestication in the Pontic–Caspian steppe by the third millennium bc, and offer strong support for the notion that the novel exploitation of secondary animal products was a key driver of the expansions of Eurasian steppe pastoralists by the Early Bronze Age.

In vitro maturation (IVM) of oocytes has still a negative impact on the developmental competence of oocytes. Therefore, this study analysed the cumulus proteome of individual cumulus-oocyte complexes (COCs) with and without maturational competence, matured under in vivo or in vitro conditions (n = 5 per group). A novel, ultrasensitive mass spectrometry (MS) based protein profiling approach, using label-free quantification, was applied. The detected cumulus proteome included 2226 quantifiable proteins and was highly influenced by the maturation condition (479 differentially expressed proteins) as well as maturational competence of the corresponding oocyte (424 differentially expressed proteins). Enrichment analysis showed an overrepresentation of the complement and coagulation cascades (CCC), ECM-receptor interaction and steroid biosynthesis in cumulus of COCs that matured successfully under in vivo conditions. Verification of the origin of CCC proteins was achieved through detection of C3 secretion into the maturation medium, with significantly increasing concentrations from 12 (48.4 ng/ ml) to 24 hours (68 ng/ml: p < 0.001). In relation, concentrations in follicular fluid, reflecting the in vivo situation, were >100x higher. In summary, this study identified important pathways that are impaired in IVM cumulus, as well as potential markers of the maturational competence of oocytes.