Answering questions about APA

BUFFALO, N.Y.—Pinpointing culprits that aid in the progression of cancer, such as oncogenes or easily hijacked immune checkpoints, improves our understanding of the disease and provides new targets for combating it. One such culprit, a gene regulatory mechanism known as alternative polyadenylation, was recently subjected to its first large-scale analysis in a single cancer type by Roswell Park Comprehensive Cancer Center researchers, who discovered that it plays a key role in the development of pancreatic cancer.

Dr. Michael Feigin, an assistant professor of oncology in the Department of Pharmacology and Therapeutics, led this work, along with postdoctoral fellow Dr. Swati Venkat and colleagues. Their results were published in Genome Research in an article titled “Alternative polyadenylation drives oncogenic gene expression in pancreatic ductal adenocarcinoma.” This research was funded in part by grants from the National Cancer Institute and the U.S. Department of Defense, as well as donations to Roswell Park.

Alternative polyadenylation regulates the expression of genes in cells—including oncogenes, genes known to drive cancer. As the authors of the Genome Research paper explain, “Alternative polyadenylation (APA) is a gene regulatory process that dictates mRNA 3’ UTR length, resulting in changes in mRNA stability and localization. APA is frequently disrupted in cancer and promotes tumorigenesis through altered expression of oncogenes and tumor suppressors.”

“Functional studies of the genes comprising the APA machinery have highlighted their relevance to tumor growth; for example, in glioblastoma, overexpression of the APA factor NUDT21 (a repressor of proximal 3’ UTR PAS usage) reduces tumor cell proliferation and inhibits tumor growth in vivo (Masamha et al. 2014). Subsequently, a number of pan-cancer analyses have utilized standard RNA-sequencing (RNA-seq) data to identify 3’ UTR shortening and lengthening events across cancer types (Le Pera et al. 2015; Feng et al. 2017; Ye et al. 2018; Grassi et al. 2016; Xia et al. 2014). While these analyses have uncovered recurrent APA events across multiple tumor types, they also detected tumor type-specific events (Xue et al. 2018). Additionally, differential 3’ UTR processing has been shown to drive tissue-specific gene expression,” they added.

The effect of oncogenes lies in the proteins they produce. Messenger RNA (mRNA) is a single-strand complement to DNA that encodes the genetic information of a cell, is “read” by ribosomes and then is “translated” to form proteins.

The length of the genetic sequence at the end of a molecule—referred to as a 3 prime untranslated region, or 3’ UTR—influences the amount of protein that is produced. When those lengths are shorter, it results in increased protein production, which in turn leads to cancerous growth. According to the authors, “Changes in 3’ UTR length can modulate mRNA stability, function or subcellular localization through disruption of miRNA or RNA-binding protein regulation.” When the researchers compared tissue samples from pancreatic cancer patients with samples from normal, healthy pancreases, they found a noted difference in 3’ UTR lengths.

“We found widespread, recurrently shorter 3’ UTR lengths of multiple oncogenes in pancreatic cancer patients,” remarked Venkat, first author on the study. “This shorter length significantly increased gene expression of known pancreatic cancer oncogenes.”

“We conclude that 3’ UTR length changes drive widespread oncogene dysregulation in pancreatic cancer,” Feigin added. “Our next step will be to unravel the protein machinery that regulates these length changes and target these proteins to explore possible therapeutic avenues.”

In addition, the authors also reported on the discovery of a novel growth-promoting gene known as casein kinase 1 alpha. This gene’s mRNA presents with a shorter 3’ UTR length in cancer cells that results in increased expression and cell growth. They noted that knockdown or inhibition of CSNK1A1, the gene that encodes for casein kinase 1 alpha, “attenuates PDA cell proliferation and clonogenic growth.”

“[Our] single-cancer approach can identify APA-regulated, disease-specific vulnerabilities. Our computational analysis and experimental validation have revealed unexpected mediators of PDA biology and broadened our understanding of the regulatory role of 3’ UTR sequence space in cancer. This comprehensive analysis reveals the scope of previously uncharacterized APA events in regulating functionally relevant PDA genes, improving patient prognosis and driving tumor evolution. We propose that the landscape of 3’ UTR alterations in PDA represents a novel avenue to better understand PDA progression and identify new drug targets,” the authors concluded.