Arginylated Calreticulin at Plasma Membrane Increases Susceptibility of Cells to Apoptosis

Abstract

Background: Calreticulin retrotranslocated from the endoplasmic reticulum to the cytoplasm is post-translationally arginylated associates to stress granules following stress that decrease Ca 2ϩ . Results: Arginylated calreticulin reaches the plasma membrane as a response to stress. Conclusion: Arginylation confers to calreticulin a function as one of the preapoptotic signals of the cells. Significance: Arginylated calreticulin is a novel factor involved in stress-induced apoptosis.

Calreticulin and Arginylated Calreticulin Have Different Susceptibilities to Proteasomal Degradation

“…What is the cause of the higher half-life of R-CRT? We demonstrated previously that R-CRT but not CRT is recruited to SGs and that R-CRT moves to the plasma membrane in response to stress, acting as a pro-apoptotic signal (11). Arginylation may, therefore, protect CRT from proteasomal degradation under stress conditions by altering its metabolic fate.…”

“…3). For cloning of pEGFP-Ub-CRT and pEGFP-Ub-R-CRT cDNAs, the coding sequences of Ub-CRT and R-CRT were amplified by PCR from pHUE-CRT-FLAG and pHUE-R-CRT-FLAG expression vectors, respectively (11). The forward primer (same for both amplification reactions) contained an EcoRI restriction site (underlined), a Kozak sequence (boldfaced), and the Ub sequence (italicized): 5Ј-CCGGAATTCCCCGCCGCCACC-ATGCAGATCTTTGTGAAGACCCTCACTGG-3Ј.…”

“…Assessment of Cell Death-Cells/sample were collected as in López Sambrooks et al (11). Briefly, cells were washed in PBS, pelleted, and resuspended in buffer (10 mM HEPES/NaOH, pH 7.4, 140 mM NaCl, 2.5 mM CaCl 2 ) containing propidium iodide to identify dead cells.…”

“…Arginylation is a PTM mediated by arginyl-tRNA-protein transferase (Ate1), which transfers Arg from arginyl-tRNA onto proteins that have an acidic amino acid or Cys as the N-terminal residue (4 -6). Many proteins are arginylated in vivo (7,8), and this PTM plays key roles in a variety of physiological processes (3) including cell adhesion (9,10), apoptosis (11), and cellular stress (7,12,13).…”

“…We showed that arginylation of CRT is essential for its association with stress granules (SGs) and promotes its dimerization (7,13). Apoptosis induction is associated with an increased R-CRT level at the cell surface (11). Surface exposure of CRT on tumor cells has been suggested to promote their uptake by phagocytes (18).…”