All tissues in the human body must be able to replace dying cells and repair wounds to maintain structural integrity and function. Regeneration is critical for the skin and, in particular, for its outermost layer, the epidermis, which is replaced every 40-56 days. Stem cells in the epidermal basal layer –called basal progenitor cells– sustain the high cellular turnover required for the epidermis’ growth and development. Basal cells maintain their population (self-renewal), create more of themselves (proliferation), and give rise to specialized cell types (differentiation). The fate of basal cells is tightly coordinated not only to maintain homeostasis and respond to wounding but also to avoid uncontrolled expansion of cancerous cells. The Beronja lab (Human Biology Division) is interested in understanding the mechanisms that regulate the balance between malignant growth restriction and essential epidermal growth.
A previous study from the Beronja lab demonstrated that the expansion of mutant cells could be limited by a mechanism that biases their fate towards differentiation, which over time leads to their elimination from the skin; however, this study only investigated the epidermis’ growth restrictive potential of mutant cells that arise from single clones. In a recent study, published in the journal Cell Stem Cell, investigators in the Beronja lab, led by MD-PhD graduate student Elise Cai, sought to identify the mechanisms that keep larger clusters of mutant cells in check while maintaining epidermal development. They discovered a translational mechanism that suppresses tumor formation by regulating stem cell fate during widespread oncogenic activation and proliferation.
The investigators first determined the effects on basal progenitor cell fate upon widespread oncogene activation using a mouse model with conditional epidermal expression of the oncogene HrasG12V. They observed an increase in the rate of basal cell proliferation in the HrasG12V epidermis. Tissue growth modeling based on proliferation rates predicted a 25-fold increase in epidermal expansion in the HrasG12V epidermis over wild-type epidermis. However, surprisingly, experimentally observed tissue expansion upon HrasG12V activation was only 4-fold higher than wild type. The authors hypothesized that the increased basal cell proliferation in HrasG12V epidermis could be counteracted by a moderate suppression of basal cell renewal to limit epidermal growth. After following the fate of progenitors in the HrasG12V epidermal basal layer and their progeny after cell division, the investigators determined that although basal cells were proliferating at an increased rate, their renewal rate was lower – a switch from the regulation observed in wild-type epidermis.
Next, the investigators set out to identify a molecular mechanism with the potential to coordinate stem cell responses in the dynamic environment of the developing epidermis. They found that protein synthesis was elevated in the HrasG12V basal progenitors and linked this finding to previous reports that describe increased protein synthesis as an important factor mediating the epidermal response to oncogenic activation. Thus, the investigators proposed that the translational apparatus could be implicated in the epidermal response following HrasG12V activation. To identify the translational machinery components that regulate HrasG12V progenitor proliferation and renewal, the researchers designed two in vivo functional screens (a proliferation and a renewal screen) using a library of short hairpin RNAs that target translational machinery genes. The investigators classified the screen hits into pro-proliferation, anti-proliferation, pro-renewal, and anti-renewal genes, and focused on those that simultaneously promoted HrasG12V progenitor proliferation and inhibited self-renewal. In the overlap, they identified eIF2B5, a gene implicated in epithelial cancers and congenital disorders involving tissue-specific defects.
In HrasG12V epidermis, eIF2B5 depletion rescued translation rate, progenitor cell proliferation rate, and renewal probability to wild-type levels. In contrast, eIF2B5 reduction in wild type cells did not affect progenitor cell behaviors, demonstrating that eIF2B5 stem cell regulatory effects are specific to widespread oncogenic activation. In vivo ribosome profiling, a deep sequencing-based method used to measure global translation in detail, revealed eIF2B5 controls the translation of a particular subset of HrasG12V-dependent mRNAs, including many cancer-related genes. To identify genes regulated by eIF2B5 that could specifically inhibit renewal and drive proliferation, the investigators performed a second short hairpin RNA screen to target eIF2B5-regulated oncogenes. This screen revealed that eIF2B5 drives the translation of different translational networks that independently regulate renewal and proliferation. A top hit from the screen, FBXO32, is a ubiquitin ligase previously implicated in both tumor suppression and oncogenesis. Further characterization in the HrasG12V epidermis revealed that FBXO32 only blocks renewal without affecting proliferation to restrain malignant overgrowth. The authors conclude that the functional segregation of proliferation and renewal upon widespread oncogenic activation could allow the epidermis to dynamically adapt stem cell fate choices without compromising its physiological need for continual proliferation.
This research was supported by grants from the National Institutes of Health
UW/Fred Hutch Cancer Consortium members Dr. Sloban Beronja and Dr. Andrew C. Hsieh contributed to this work.
Cai EY, Kufeld MN, Schuster S, Arora S, Larkin M, Germanos AA, Hsieh AC, Beronja S. (2020). Selective Translation of Cell Fate Regulators Mediates Tolerance to Broad Oncogenic Stress. Cell Stem Cell. DOI: 10.1016/j.stem.2020.05.007