Acylpeptide hydrolase is a novel regulator of KRAS plasma membrane localization and function

Abstract

The primary site for KRAS signaling is the inner leaflet of the plasma membrane (PM). We previously reported that oxanthroquinone G01 (G01) inhibited KRAS PM localization and blocked KRAS signaling. In this study, we identified acylpeptide hydrolase (APEH) as a molecular target of G01. APEH formed a stable complex with biotinylated G01, and the enzymatic activity of APEH was inhibited by G01. APEH knockdown caused profound mislocalization of KRAS and reduced clustering of KRAS that remained PM localized. APEH knockdown also disrupted the PM localization of phosphatidylserine (PtdSer), a lipid critical for KRAS PM binding and clustering. The mislocalization of KRAS was fully rescued by ectopic expression of APEH in knockdown cells. APEH knockdown disrupted the endocytic recycling of epidermal growth factor receptor and transferrin receptor, suggesting that abrogation of recycling endosome function was mechanistically linked to the loss of KRAS and PtdSer from the PM. APEH knockdown abrogated RAS-RAF-MAPK signaling in cells expressing the constitutively active (oncogenic) mutant of KRAS (KRASG12V), and selectively inhibited the proliferation of KRAStransformed pancreatic cancer cells. Taken together, these results identify APEH as a novel drug target for a potential anti-KRAS therapeutic.

Precise and efficient silencing of mutant KrasG12D by CRISPR-CasRx controls pancreatic cancer progression

“…Consistent with the above data, CasRx-gRNA resulted in a dramatic reduction of Kras protein in PANC-1 cells (Figure 2 A-B) but not in H6c7 or MIAPaCa-2 cells (Figure 2 A-B, Figure S3 B). To recruit and activate downstream signaling pathways, Kras must localize primarily to the inner leaflet of the plasma membrane (PM) 19 . Compared to the control, we observed an obvious reduction of accumulated Kras protein near PM in CasRx + /gRNA + PANC-1 (Figure 2 C, Figure S2 A-B).…”

RAS: Striking at the Core of the Oncogenic Circuitry

“…RAS acetylation has also been shown to reduce signaling activity, with cells dependent on the protein deacetylases HDAC6 and SIRT2 to maintain RAS signaling (26, 27). Additionally, acylpeptide hydrolase (APEH) contributes to the appropriate localization of RAS to the plasma membrane by regulating phosphatidylserines in the plasma membrane (28) (Figure 1).…”

The KRAS and other prenylated polybasic domain membrane anchors recognize phosphatidylserine acyl chain structure

“…In consequence, PtdSer levels in the PM modulate the extent of KRAS localization to, and nanoclustering on, the PM and hence regulate KRAS-dependent effector recruitment and signaling (7,11,14). Depletion of PtdSer compromises the proliferation of KRAS-driven cancer cell lines and KRAS oncogenicity in mouse xenograft models (15)(16)(17)(18)(19)(20). Thus, PtdSer is a key structural component of KRAS signaling nanoclusters on the PM and plays important roles in KRAS function and pathology.…”