# The translation regulation of pro-apoptotic genes

> **NIH NIH R01** · OHIO STATE UNIVERSITY · 2022 · $86,023

## Abstract

PROJECT SUMMARY (PARENTAL AWARD)
Inactivation of the Retinoblastoma 1 protein (pRB) is found in a substantial fraction of human cancers. This is
because pRB inhibits the activity of three important E2F transcription factors (E2F1-3) that control the expression
of numerous tumor-promoting genes, which control cell growth including cell cycle genes and metabolic factors.
Conversely, the E2Fs also regulate the transcription of tumor-inhibiting processes including important apoptosis
and necrosis genes. This Rb-controlled coupled expression of proliferation and cell death genes is designed to
prevent uncontrolled cell growth that is a hallmark of cancer. However, the widespread loss of pRB in cancer
underlines a major unsolved puzzle: why don’t pRB-deficient cells simply die?
To answer this question, we profiled RNA and protein changes following pRB-depletion and found that essential
cell death genes were indeed transcribed, but the mRNAs were not translated into functional proteins. We then
searched for RNA-binding proteins (RBPs) that bound to and blocked the Ribosome occupancy of these cell-
death genes. This analysis identified the PUMILIO (PUM) family of RBPs as potential regulators of cell death
RNAs. Importantly, as would be predicted by such a regulatory role, co-depletion of PUM1 and PUM2 resulted
in the apoptosis of RB1-deficient cancer cells. However, the high level of functional redundancy has limited
functional dissection of PUM1 and PUM2 mechanics in specific translational regulation of cell death mRNAs. To
circumvent this technical issue, we have engineered PUM1-/-, PUM2-/- CRISPR-derived knockout human cells
that additionally express individually degradable PUM proteins. Following the addition of the plant hormone,
AUXIN, PUM1 or PUM2 are rapidly degraded, enabling kinetic analysis of the stability, localization and
translation initiation of Rb-regulated mRNA transcripts. Using well-defined candidate cell death genes, we
propose to determine the mechanism(s) associated with mRNA regulation by the individual PUM1 or PUM2
proteins. We will then determine which PUM regulatory mechanisms are required in the development of RB1-/-
Small Cell Lung Cancer (SCLC) in vivo.

## Key facts

- **NIH application ID:** 10599664
- **Project number:** 3R01CA251753-03S1
- **Recipient organization:** OHIO STATE UNIVERSITY
- **Principal Investigator:** Wayne Miles
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $86,023
- **Award type:** 3
- **Project period:** 2020-07-14 → 2025-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10599664

## Citation

> US National Institutes of Health, RePORTER application 10599664, The translation regulation of pro-apoptotic genes (3R01CA251753-03S1). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10599664. Licensed CC0.

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