SINC a new type III secreted protein of the avian and human pathogen uniquely targets the nuclear envelope of are ubiquitous Gram-negative bacteria capable of infecting a wide variety of hosts and tissues (Corsaro and Venditti 2004 ). (Stamm 1999 ). Two other species and is widespread-detected in nearly 500 avian species (Kaleta and Taday 2003 )-in which infection can be either latent or systemic with clinically overt respiratory symptoms (Stewardson and Grayson 2010 ). is a highly infectious medically significant potential human pathogen classified as a category B bioterrorism agent by the Centers for Disease Control and Prevention (www.bt.cdc.gov/agent/agentlist-category.asp). Inhalation via aerosols can cause life-threatening pneumonia (Smith to be significantly more infectious and pathogenic than in humans are not understood. All species are obligate intracellular pathogens with a unique developmental life cycle involving two cellular forms. After entering the host cell via endocytosis metabolically dormant chlamydiae-termed elementary bodies (EBs)-differentiate into larger actively replicating reticulate bodies (RBs) within a membrane-bound vacuole termed the inclusion. RBs differentiate back into EBs asynchronously so the chlamydial inclusion includes both forms (RBs and EBs) at late stages of infection. After completing development EBs exit upon lysis of the host cell or nonexocytic extrusion of whole or part of the inclusion (Hybiske and Stephens 2007 ) and then either disseminate or infect neighboring cells. All species encode a complete type III secretion (T3S) system that enables the direct translocation of effector proteins across both the bacterial envelope and host plasma membrane-derived inclusion membrane into the host cytosol where they target specific host proteins and pathways to promote and maintain infection (Peters inclusion surface (Derre YopN (Fields and Hackstadt 2000 ) but also modulates the host cytoskeleton (Archuleta (Wang and the impracticality of clonal isolation. Strategies that have been successful include identification based on homology to effectors from other Rabbit polyclonal to ZNF703.Zinc-finger proteins contain DNA-binding domains and have a wide variety of functions, most ofwhich encompass some form of transcriptional activation or repression. ZNF703 (zinc fingerprotein 703) is a 590 amino acid nuclear protein that contains one C2H2-type zinc finger and isthought to play a role in transcriptional regulation. Multiple isoforms of ZNF703 exist due toalternative splicing events. The gene encoding ZNF703 maps to human chromosome 8, whichconsists of nearly 146 million base pairs, houses more than 800 genes and is associated with avariety of diseases and malignancies. Schizophrenia, bipolar disorder, Trisomy 8, Pfeiffer syndrome,congenital hypothyroidism, Waardenburg syndrome and some leukemias and lymphomas arethought to occur as a result of defects in specific genes that map to chromosome 8. bacterial genera (Hsia (Fields and Hackstadt 2000 ; Subtil as a surrogate to test putative chlamydial T3S-dependent secreted proteins predicted by the protein homology-based algorithm SIEVE (Samudrala (NCBI G5Q_0070) of strain CAL10 as a putative effector (Hovis protein (SINC) based on its novel localization at the nuclear envelope (NE) of infected and neighboring uninfected cells and association with nuclear membrane proteins. RESULTS is syntenic and encodes a weak orthologue of CT694 The putative 25-hydroxy Cholesterol effector gene was chosen for further investigation because it posed a paradox: is syntenic with of each downstream of the phosphoglycerate kinase gene (Supplemental Figure S1A); however the encoded CT694 and SINC proteins are only 12.5% identical compared 25-hydroxy Cholesterol with 74% identical phosphoglycerate kinase proteins. Residual identity to CT694 is scattered throughout SINC (e.g. residues 1-11 151 and 458-466) suggesting divergence from a common ancestral gene. Low sequence identity suggested that SINC and CT694 were functionally distinct and might therefore be expressed at different stages 25-hydroxy Cholesterol of development in or CAL10 revealed low or background levels of transcripts from 6 to 24 h postinfection (hpi) peaking at 30-42 hpi and decreasing sharply by 42 hpi with a strong trend toward statistical significance (= 13.675 = 0.057; Supplemental Figure S1B) similar to and and their gene products were expressed at similar times during development (Belland CAL10-infected HeLa cells fixed with methanol at 24 hpi and stained using antibodies specific for SINC … FIGURE 2: SINC is secreted by chlamydiae and targets the nuclear envelope of infected and uninfected neighboring cells late 25-hydroxy Cholesterol in development. Immunofluorescence images of CAL10-infected HeLa cells fixed with methanol at 36 hpi and stained using … At 36 hpi nearly all chlamydiae within the inclusion were SINC positive as visualized by confocal microscopy (Figure 2A). We also detected strong SINC-specific fluorescence at the host cell NE especially on the side nearest the inclusion (Figure 2A) and weak SINC staining in the nucleoplasm (Figure 2B) consistent with IEM (Figure 1C). These and later images hinted that SINC might colocalize with pore-linked filaments extending into the nucleoplasm (e.g. white arrowheads in Figures 1C and ?and3A;3A; Arlucea CAL10-infected.