Paper

Alt-RPL36 downregulates the PI3K-AKT-mTOR signaling pathway by interacting with TMEM24

Na, Zhenkun, Cao, Xiongwen, Uttamapinant, Chayasith, Olatunji, Elizabeth, Sappakhaw, Khomkrit, Luo, Yang, Phoodokmai, Thitima, Slavoff, Sarah A, Khitun, Alexandra

2020

Overview

Abstract Thousands of previously unannotated small and alternative open reading frames (alt-ORFs) have recently been revealed in the human genome, and hundreds are now known to be required for cell proliferation. Many alt-ORFs are co-encoded with proteins of known function in multicistronic human genes, but the functions of only a handful are currently known in molecular detail. Using a proteomic strategy for discovery of unannotated short open reading frames in human cells, we report the detection of alt-RPL36, a 148-amino acid protein co-encoded with and overlapping human RPL36 (ribosomal protein L36). Alt-RPL36 partially localizes to the endoplasmic reticulum, where it interacts with TMEM24, which transports the phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] precursor phosphatidylinositol from the endoplasmic reticulum to the plasma membrane. Knock-out of alt-RPL36 in HEK 293T cells increases PI(4,5)P2 levels in the plasma membrane, upregulates the PI3K-AKT-mTOR signaling pathway, and increases cell size. Four serine residues of alt-RPL36 are phosphorylated, and mutation of these four serines to alanine abolishes interaction with TMEM24 and, consequently, abolishes alt-RPL36 effects on PI3K signaling and cell size. These results implicate alt-RPL36 as a novel regulator of PI(4,5)P2 synthesis upstream of the PI3K-AKT-mTOR signaling pathway. More broadly, these results show that the alt-RPL36 transcript can express two sequence-independent polypeptides from overlapping ORFs that regulate the same process – protein synthesis – via different molecular mechanisms (PI3K signaling and ribosome composition), expanding our knowledge of the mechanisms by which multicistronic human genes function. Competing Interest Statement The authors have declared no competing interest. Footnotes * Figures 2, 6, 7 and 8 revised. Supplemental files updated.

• Publisher: Cold Spring Harbor Laboratory Press,Cold Spring Harbor Laboratory
• Subject: 1-Phosphatidylinositol 3-kinase / AKT protein / Alanine / Endoplasmic reticulum / Genomes / Genomics / Kinases / Molecular modelling / Open reading frames / Phosphatidylinositol 4,5-diphosphate / Post-translation / Protein biosynthesis / Proteins / Proteomics / Serine / Signal transduction / TOR protein / Transcription