Ral GTPase–activating protein regulates the malignancy of pancreatic ductal adenocarcinoma

Abstract

The small GTPases RalA and RalB are members of the Ras family and activated downstream of Ras. Ral proteins are found in GTP‐bound active and GDP‐bound inactive forms. The activation process is executed by guanine nucleotide exchange factors, while inactivation is mediated by GTPase‐activating proteins (GAPs). RalGAPs are complexes that consist of a catalytic α1 or α2 subunit together with a common β subunit. Several reports implicate the importance of Ral in pancreatic ductal adenocarcinoma (PDAC). However, there are few reports on the relationship between levels of RalGAP expression and malignancy in PDAC. We generated RalGAPβ‐deficient PDAC cells by CRISPR‐Cas9 genome editing to investigate how increased Ral activity affects malignant phenotypes of PDAC cells. RalGAPβ‐deficient PDAC cells exhibited several‐fold higher Ral activity relative to control cells. They had a high migratory and invasive capacity. The RalGAPβ‐deficient cells grew more rapidly than control cells when injected subcutaneously into nude mice. When injected into the spleen, the RalGAPβ‐deficient cells formed larger splenic tumors with more liver metastases, and unlike controls, they disseminated into the abdominal cavity. These results indicate that RalGAPβ deficiency in PDAC cells contributes to high activities of RalA and RalB, leading to enhanced cell migration and invasion in vitro, and tumor growth and metastasis in vivo.

Genomic analysis reveals HDAC1 regulates clinically relevant transcriptional programs in pancreatic cancer

“…We used biochemical assays to confirm that MP2_HDAC1_OE cells have higher GTPase activity than control cells and that cells treated with a HDACi have further reduced GTPase activity. GTPase signaling is involved in pancreatic cancer initiation, metastasis, and drug resistance 46 . Increased GTPase signaling leads to the activation of key pathways, such as MAPK and PI3K, that regulate cell proliferation, migration, and survival in cancer 26 .…”

“…Throughout this study, our gene set enrichment analyses of genes associated with HDAC1 overexpression consistently revealed GTPase activity as an enriched process. A variety of proteins contribute to GTPase activity and many are druggable 47 , which makes them of potential clinical interest. We have shown that expression of GTPases and GTPase activating proteins are associated with worse overall survival.…”

“…This effect was reversible in cells treated with a HDACi, which reduced GTPase activity. GTPase signaling is important for pancreatic cancer initiation, metastasis, and invasion 48 . Increased GTPase signaling leads to the activation of key signaling cascades, such as MAPK and PI3K, that regulate cell proliferation, migration, and survival in cancer 27 .…”

Ral GTPase promotes metastasis of pancreatic ductal adenocarcinoma via elevation of TGF-β1 production

“…Most tissues express both α subunits, with α1 predominant in the brain and α2 in the liver and lung ( 8 ). In the pancreas, the expression levels of α1 and α2 subunits are similar ( 9 ).…”

“…In pancreatic cancer, Ral has been shown to be highly activated in cancer tissues with K-Ras mutations ( 18 ). We have previously generated RalGAPβ-deficient PDAC cells using two cell lines, MIA PaCa-2 and PANC-1, and found that these cells have high Ral activity and exhibit high invasive and metastatic capacities ( 9 ). However, since Ral can interact with various downstream effectors such as the exocyst complex, RalBP1, phospholipase D, and M-Sec, it remains unclear how Ral regulates invasion and metastasis of PDAC cells.…”