[F31] Identifying novel regulators of KRAS translation
Ente: National Cancer Institute
Scadenza: 2029-06-30
Importo max: 49.612 EUR
Paese: US
Descrizione
PROJECT ABSTRACT:
KRAS is a central oncogenic driver of pancreatic ductal adenocarcinoma (PDAC), the third leading cause of
cancer mortality in the United States. KRAS dosage is critical to driving tumorigenesis; however, how oncogenes
are translationally regulated to drive dosage remains poorly understood. The RNA levels of key oncogenic drivers
often do not correlate with protein abundance, emphasizing the importance of translational control. The 5´
untranslated regions (UTR) of oncogenic mRNAs possess cis-elements, such as specific sequences and
structures that, in normal cells, keep translation tightly regulated to maintain accurate protein dosage. However,
this process is broken in cancer. KRAS is a key example of this, as it possesses a highly structured 5´UTR, yet
which specific RNA binding proteins (RBP) interact with its 5´UTR and how they influence its translation
efficiency, as well as how cancer cells exploit this process to promote KRAS dosage, are all areas that are still
largely unexplored. To address this outstanding question, we used a selective translational reporter coupled with
genome-wide CRISPRi screening and fluorescence-activated cell sorting to identify regulators of KRAS
translation through its 5´UTR in human PDAC cells. For this proposal, we are exploring the top two activators
identified in the screen: Upstream of N-Ras (UNR) and its interacting protein (UNRIP). UNR is an RBP that
bridges UNRIP with target mRNAs; however, little is known about the mechanism of action and neither have
been previously reported to regulate the KRAS mRNA or PDAC. I hypothesize that UNR and UNRIP promote
KRAS translation through the 5´UTR to favor PDAC cell growth and survival. We found that the depletion of UNR
and UNRIP decreases the translation and protein levels of KRAS. In Aim 1, we propose to determine the
mechanism by which UNR and UNRIP promote KRAS translation through the 5´UTR using polysome profiling,
luciferase assays, translation initiation complex formation assays, and cross-linked immunoprecipitation
sequencing. In Aim 2, we will define the role of UNR and UNRIP in driving PDAC through KRAS translation in
vitro and in vivo. We found a decrease in the colony formation capacity of PDAC cells upon UNR and UNRIP
depletion, which was dependent upon KRAS expression. To further explore these compelling results, we will use
a range of cell viability assays across various PDAC models that range in KRAS dependency. Additionally, we
will test the therapeutic relevance of this regulation using RAS signaling inhibitors and in vivo mouse models.
Altogether, this project will determine how PDAC coordinates the selective translation of KRAS through a novel
mechanism of post-transcriptional gene regulation, ultimately leading to the discovery of selective avenues for
therapeutic intervention in cancer. The proposed research will be conducted in the scientifically rich environment
of UCSF under the mentorship of an expert in the field. There i
Istituzione: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
PI: Kenya Bonitto
Progetto: 1F31CA310485-01A1
Settori: National Cancer Institute
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