Trafficking and proteolytic processing of RNF13, a model PA-TM-RING family endosomal membrane ubiquitin ligase.

RING finger protein 13 (RNF13) is a ubiquitously expressed, highly regulated ubiquitin ligase anchored in endosome membranes. A RING domain located in the cytoplasmic half of this type 1 membrane protein mediates ubiquitination in vitro but physiological substrates have not yet been identified. The protein localized in endosomal membranes undergoes extensive proteolysis in a proteasome-dependent manner, but the mRNA level can be increased and the encoded protein stabilized under specific physiological conditions.

Nuclear targeting of an endosomal E3 ubiquitin ligase.

Ring finger protein 13 (RNF13) is an E3 ubiquitin ligase embedded in endosome membranes. The protein undergoes constitutive post-translational proteolysis, making its detection difficult unless cells are incubated with a proteasome inhibitor to allow biosynthetic forms to accumulate. When cells were treated with phorbol 12-myristate 13-acetate (PMA), RNF13 avoided proteolysis.

The PA-TM-RING protein RING finger protein 13 is an endosomal integral membrane E3 ubiquitin ligase whose RING finger domain is released to the cytoplasm by proteolysis.

PA-TM-RING proteins have an N-terminal protease-associated domain, a structure found in numerous proteases and implicated in protein binding, and C-terminal RING finger and PEST domains. Homologous proteins include GRAIL (gene related to anergy in leukocytes), which controls T-cell anergy, and AtRMR1 (receptor homology region-transmembrane domain-RING-H2 motif protein), a plant protein storage vacuole sorting receptor. Another family member, chicken RING zinc finger (C-RZF), was identified as being upregulated in embryonic chicken brain cells grown in the presence of tenascin-C.

Factor H binding to PspC of Streptococcus pneumoniae increases adherence to human cell lines in vitro and enhances invasion of mouse lungs in vivo.

Pneumococcal surface protein C (PspC) binds to both human secretory immunoglobulin A (sIgA) and complement factor H (FH). FH, a regulator of the alternative pathway of complement, can also mediate adherence of different host cells. Since PspC contributes to adherence and invasion of host cells, we hypothesized that the interaction of PspC with FH may also mediate adherence of pneumococci to human cells.

Interaction of clinical isolates of Streptococcus pneumoniae with human complement factor H.

PspC recruits complement factor H (FH) to the pneumococcal surface. While there is differential expression of pspC during infection, detection of PspC on the surface of viable pneumococci is difficult due to variability among PspCs. We analyzed FH binding to detect PspC expression on the surface of pneumococcal isolates from different pathological sources.

Antigen three-dimensional structure guides the processing and presentation of helper T-cell epitopes.

Antigen three-dimensional structure potentially controls presentation of CD4(+) T-cell epitopes by limiting the access of proteolytic enzymes and MHC class II antigen-presenting proteins. The protease-sensitive mobile loops of Hsp10s from mycobacteria, Escherichia coli, and bacteriophage T4 (T4Hsp10) are associated with adjacent immunodominant helper T-cell epitopes, and a mobile-loop deletion in T4Hsp10 eliminated the protease sensitivity and the associated epitope immunodominance. In the present work, protease-sensitivity and epitope presentation was analyzed in a group of T4Hsp10 variants.

In vivo binding of complement regulator factor H by Streptococcus pneumoniae.

Pneumococcal surface protein C (PspC) binds to the complement regulatory protein factor H (FH), which inhibits alternative pathway activation. In the present study, using a mouse model of systemic infection and flow-cytometric analyses, we demonstrated an in vivo interaction between FH and pneumococci and showed differential FH binding during bacteremia. Flow-cytometric analyses of pneumococci harvested after intraperitoneal (ip) challenge demonstrated increased binding of FH, compared with that after intravenous (iv) challenge.

Dual roles of PspC, a surface protein of Streptococcus pneumoniae, in binding human secretory IgA and factor H.

Streptococcus pneumoniae, also known as the pneumococcus, contains several surface proteins that along with the polysaccharide capsule function in antiphagocytic activities and evasion of the host immune system. These pneumococcal proteins interact with the host immune system in various ways and possess a wide range of biological activities that suggests that they may be involved at different stages of pneumococcal infection. PspC, also known as CbpA and SpsA, is one of several pneumococcal surface proteins that binds host proteins, including factor H (FH) and secretory IgA (sIgA) via the secretory component.

Proteolytic sensitivity and helper T-cell epitope immunodominance associated with the mobile loop in Hsp10s.

Antigen three-dimensional structure potentially limits antigen processing and presentation to helper T-cell epitopes. The association of helper T-cell epitopes with the mobile loop in Hsp10s from mycobacteria and bacteriophage T4 suggests that the mobile loop facilitates proteolytic processing and presentation of adjacent sequences. Sites of initial proteolytic cleavage were mapped in divergent Hsp10s after treatment with a variety of proteases including cathepsin S.

Structural basis for helper T-cell and antibody epitope immunodominance in bacteriophage T4 Hsp10. Role of disordered loops.

Antigen three-dimensional structure potentially limits the access of endoproteolytic processing enzymes to cleavage sites and of class II major histocompatibility antigen-presenting proteins to helper T-cell epitopes. Helper T-cell epitopes in bacteriophage T4 Hsp10 have been mapped by restimulation of splenocytes from CBA/J and C57BL/6J mice immunized in conjunction with mutant (R192G) heat-labile enterotoxin from Escherichia coli. Promiscuously immunogenic sequences were associated with unstable loops in the three-dimensional structure of T4 Hsp10.