We decided to characterize expression of IRIP in the mouse model of LPS-induced endotoxemia. Endotoxemia can induce multiple organ ischemia due to microvascular constriction (9). In normal kidney, IRIP was expressed at a basal level and it was activated 1.7-, 4.2-, and 5.1-fold after ischemia and at 6 and 24 h of reperfusion, respectively (Fig. Therefore, we examined the expression of IRIP mRNA in mouse kidney at different time points after I/R. We identified the IRIP gene as an I/R-inducible gene in a differential display analysis. Testis and thyroid were also the tissues with the highest expression level of human IRIP, according to the SymAtlas database. This result is in excel- lent agreement with microarray data from SymAtlas database (Genomics Institute of the Novartis Research Foundation ). IRIP mRNA was expressed at a low level in the spleen, muscle, heart, and small intestine and at a relatively high level in testis, thyroid, ovary, colon, kidney, and brain. A single IRIP transcript was detected with an estimated size of 1.4 kb, which matched the size of cloned cDNA (Fig. The expression of IRIP mRNA in mouse tissues was examined by Northern blot analysis. A number of amino acid residues are identical in all these three proteins (Fig. However, several of the highly conserved sequence blocks in IRIP homologues are also conserved in SUA5 and YrdC, suggesting that these proteins probably have a common ancestry and may have diverged during evolution. The overall sequence similarities between the three proteins are quite modest (approximately 25 to 30% identity with many mismatches and large gaps), which is probably the reason why SUA5 and YrdC were not identified in the initial BLAST database search. A multiple sequence alignment of human IRIP, SUA5, and YrdC is shown in Fig. The founding members of this protein family include SUA5 ( Saccharomyces cerevisiae ) and YrdC ( E. It contains an average of about 180 amino acid residues. The SUA5/yciO/yrdC domain is also found in many prokaryotic proteins of unknown functions. PS00107) and SUA5/yciO/yrdC family signature (PROSITE accession no. Two sequence motifs were identified with a significant P value: a protein kinase ATP- binding signature (PROSITE accession no. Human IRIP sequence was scanned against protein motif databases PROSITE, Pfam, and COG using several web-based programs.
IRIP does not contain transmembrane domains and appears to be a com- pact globular protein (The PredictProtein server. Human IRIP has a predicted molecular mass of 28.3 kDa, and mouse IRIP has a molecular mass of 27.8 kDa. The GenBank accession numbers of IRIP cDNA sequences are AY283537 (mouse) and AY286019/AY286020 (human). The human IRIP gene is located on chromosome 1 and the mouse IRIP gene is located on chromosome 4. Two cDNA sequences with alternative polyadenylation sites were identified upon analysis of hIRIP transcripts in human cell lines (see next section). The chromosomal regions of both genes span about 5 kb (Fig. In this review, we elaborate on reported methods and discuss recent advances and shortcomings in this area of tracking bacterial effector translocation. Recently, the existing toolset has been expanded by newly developed state-of-the art methods to monitor bacterial effector translocation and dynamics. Various approaches have been developed to understand timing and order of effector translocation, quantities of translocated effectors and their subcellular localization upon translocation into host cells.
A comprehensive understanding of effector translocation in a spatio-temporal manner is of critical importance to gain insights into an effector’s mode of action. These effectors are translocated into host cells by means of dedicated secretion systems such as the type 3 secretion system (T3SS).
Bacteria-host interactions are characterized by the delivery of bacterial virulence factors, i.e., effectors, into host cells where they counteract host immunity and exploit host responses allowing bacterial survival and spreading.
0 kommentar(er)
