M.Sc Thesis

M.Sc StudentSukenik Sigalit
SubjectCharacterizing the Cellular Role of AIRAP in Proteasomal
DepartmentDepartment of Medicine
Supervisor DR. Ariel Stanhill
Full Thesis textFull thesis text - English Version


Arsenite (As(III)) is an environmental toxin, with harmful effects resulting from inhibition of protein function, by binding reduced vicinal sulfhydryl groups, commonly pairs of thiols in close proximity. Arsenite was shown to inhibit the ubiquitin-dependent proteolytic pathway and creates a mismatch between the load of misfolded proteins and the chaperones available to bind them. To this end, several cellular responses are stimulated, such as the induction of heat-shock genes and the Integrated Stress Response (ISR). AIRAP (Arsenite Inducible Regulatory Particle Associated Protein) is unique among known arsenite-induced genes, based on the regulation of its expression and the potential activities of its gene product. AIRAP is a 171-amino acid peptide with two conserved AN1-type zinc fingers motifs, containing cysteine-histidine rich repeats. When cells are treated with arsenite, AIRAP can be found in a complex with the 26S proteasome. Knock-out of AIRAP in cells indicated an impaired clarification of poly-ubiquitinated substrates upon acute arsenite exposure. Furthermore, the proteasome containing AIRAP showed increase peptide hydrolysis activity, a feature that can attest to conformational changes the proteasome undergoes upon AIRAP binding. Based on these results it was concluded that AIRAP is a proteasome adaptor that modified the proteasome activity to better cope with the toxic effects of arsenite. Here we show that AIRAP has high cysteine content to directly bind arsenite and thus relieves proteasome inhibition caused by arsenite. We further show that the metal-binding protein Rpn11/PSMD14, that is the proteasome De-UBiquitinating enzyme (DUB), is inhibited by arsenite. We show that AIRAP is able to relieve arsenite inflicted proteasomal inhibition and propose a model to explain the mechanistic role of AIRAP in proteasomal adaptation to protein misfolding.