Phylogenomics reveals an anomalous distribution of USP genes in metazoans.

Members of the universal stress protein (USP) family were originally identified in stressed bacteria on the basis of a shared domain, which has since been reported in a phylogenetically diverse range of prokaryotes, fungi, protists, and plants. Although not previously characterized in metazoans, here we report that USP genes are distributed in animal genomes in a unique pattern that reflects frequent independent losses and independent expansions. Multiple USP loci are present in urochordates as well as all Cnidaria and Lophotrochozoa examined, but none were detected in any of the available ecdysozoan or non-urochordate deuterostome genome data. The vast majority of the metazoan USPs are short, single-domain proteins and are phylogenetically distinct from the prokaryotic, plant, protist, and fungal members of the protein family. Whereas most of the metazoan USP genes contain introns, with few exceptions those in the cnidarian Hydra are intronless and cluster together in phylogenetic analyses. Expression patterns were determined for several cnidarian USPs, including two genes belonging to the intronless clade, and these imply diverse functions. The apparent paradox of implied diversity of roles despite high overall levels of sequence (and implied structural) similarity parallels the situation in bacteria. The absence of USP genes in ecdysozoans and most deuterostomes may be a consequence of functional redundancy or specialization in taxon-specific roles.

Analysis of the dorsal spinal cord synaptic architecture by combined proteome analysis and in situ hybridization.

The proteomic analysis of tissue samples is an analytical challenge, because identified gene products not only have to be assigned to subcellular structures, but also to cell subpopulations. We here report a strategy of combined subcellular proteomic profiling and in situ hybridization to assign proteins to subcellular sites in subsets of cells within the dorsal region of rat spinal cord. With a focus on synaptic membranes, which represent a complex membrane protein structure composed of multiple integral membrane proteins and networks of accessory structural proteins, we also compared different two-dimensional gel electrophoresis systems for the separation of the proteins. Using MALDI mass spectrometric protein identification based on peptide mass fingerprints, we identified in total 122 different gene products within the different synaptic membrane subfractions. The tissue structure of the dorsal region of the spinal cord is complex, and different layers of neurons can be distinguished neuroanatomically. Proteomic data combined with an in situ hybridization analysis for the detection of mRNA was used to assign selected gene products, namely the optical atrophy protein OPA-1, the presynaptic cytomatrix protein KIAA0378/CAST1, and the uncharacterized coiled-coil-helix-coiled-coil-helix domain containing protein 3 (hypothetical protein FLJ20420), to cell subsets of the dorsal area of the spinal cord. Most striking, KIAA0378/CAST1 mRNA was found only sparsely within the dorsal horn of the spinal cord, but highly abundant within the dorsal root ganglion. This finding, combined with the identification of KIAA0378/CAST1 within the synaptic membrane fraction of the spinal cord at the protein level, are consistent with the reported presynaptic localization of CAST, predominantly within the tissue we investigated primarily attributable to primary afferent sensory neurons. Our approach may be of use in broader studies to characterize the proteomes of neural tissue.

Signalling by the FGFR-like tyrosine kinase, Kringelchen, is essential for bud detachment in Hydra vulgaris.

Signalling through fibroblast growth factors (FGFR) is essential for proper morphogenesis in higher evolved triploblastic organisms. By screening for genes induced during morphogenesis in the diploblastic Hydra, we identified a receptor tyrosine kinase (kringelchen) with high similarity to FGFR tyrosine kinases. The gene is dynamically upregulated during budding, the asexual propagation of Hydra. Activation occurs in body regions, in which the intrinsic positional value changes. During tissue displacement in the early bud, kringelchen RNA is transiently present ubiquitously. A few hours later - coincident with the acquisition of organiser properties by the bud tip - a few cells in the apical tip express the gene strongly. About 20 hours after the onset of evagination, expression is switched on in a ring of cells surrounding the bud base, and shortly thereafter vanishes from the apical expression zone. The basal ring persists in the parent during tissue contraction and foot formation in the young polyp, until several hours after bud detachment. Inhibition of bud detachment by head regeneration results in severe distortion, disruption or even complete loss of the well-defined ring-like expression zone. Inhibition of FGFR signalling by SU5402 or, alternatively, inhibition of translation by phosphorothioate antisense oligonucleotides inhibited detachment of buds, indicating that, despite the dynamic expression pattern, the crucial phase for FGFR signalling in Hydra morphogenesis lies in bud detachment. Although Kringelchen groups with the FGFR family, it is not known whether this protein is able to bind FGFs, which have not been isolated from Hydra so far.