Biology subjects

Klein, P. S.

Publications and source records attributed to Klein, P. S..

4 recordsLinked to original sources

bioRxiv2025

APC loss promotes intestinal transformation through induction of bistable stem cell states

Colorectal cancer (CRC) is a leading cause of cancer deaths, predominantly initiated by genetic inactivation of the APC tumor suppressor. Current dogma holds that APC inactivation promotes tumorigenesis via hyperactivation of the WNT pathway transcriptional effector, {beta}-CATENIN. Although {beta}-CATENIN activation is required for tumor initiation, activating mutations in {beta}-CATENIN are infrequent. Here, we ask what underlies the selective pressure for APC inactivation by comparing the oncogenic effects of APC loss to {beta}-CATENIN hyperactivation. We find that APC loss activates a {beta}-CATENIN-independent fetal intestinal transcriptional program driven by dysregulation of GSK-3 activity upon the LIM-domain protein AJUBA, a positive regulator of YAP. This results in AJUBA stabilization and downstream transcriptional activation of the YAP-driven fetal intestinal gene expression program. We find that {beta}-CATENIN- and YAP-driven transcriptional states are mutually exclusive, existing in an interchangeable bistable balance among APC-null cells. This results in more robust tumor initiation and metastatic progression downstream of APC loss relative to {beta}-CATENIN activation. Taken together, our findings explain the preferential selection for APC inactivation in CRC development and illuminate how {beta}-CATENIN- and YAP-driven gene expression programs coexist to promote tumorigenesis.

cancer biology↗

bioRxiv2023

Expansion of human hematopoietic stem cells by inhibiting translation

Hematopoietic stem cell (HSC) transplantation using umbilical cord blood (UCB) is a potentially life-saving treatment for leukemia and bone marrow failure but is limited by the low number of HSCs in UCB. The loss of HSCs after ex vivo manipulation is also a major obstacle to gene editing for inherited blood disorders. HSCs require a low rate of translation to maintain their capacity for self-renewal, but hematopoietic cytokines used to expand HSCs stimulate protein synthesis and impair long-term self-renewal. We previously described cytokine-free conditions that maintain but do not expand human and mouse HSCs ex vivo. Here we performed a high throughput screen and identified translation inhibitors that allow ex vivo expansion of human HSCs while minimizing cytokine exposure. Transplantation assays show a [~]5-fold expansion of long-term HSCs from UCB after one week of culture in low cytokine conditions. Single cell transcriptomic analysis demonstrates maintenance of HSCs expressing mediators of the unfolded protein stress response, further supporting the importance of regulated proteostasis in HSC maintenance and expansion. These culture conditions maintain adult HSCs after CRISPR/Cas9 editing of the BCL11A+58 enhancer, overcoming a major obstacle to ex vivo gene correction for human hemoglobinopathies. SignificanceO_LIWe report ex vivo expansion of human hematopoietic stem cells from umbilical cord blood using low cytokines and small molecule inhibitors of GSK-3, mTORC1, and the translation initiation factor eIF4E. C_LIO_LIOur method maintains human HSCs ex vivo after gene editing with conventional CRISPR/Cas9, lipid nanoparticle delivery, and base editing approaches, overcoming a major obstacle to gene editing for inherited blood disorders. C_LI

cell biology↗

bioRxiv2023

Targeting mitophagy in SRSF2 mutant hematologic malignancies

Splicing factor mutations are common in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML), but how they alter cellular functions is unclear. We show that the pathogenic SRSF2P95H/+ mutation disrupts the splicing of mitochondrial mRNAs, impairs mitochondrial complex I function, and robustly increases mitophagy. We also identified a mitochondrial surveillance mechanism by which mitochondrial dysfunction modifies splicing of the mitophagy activator PINK1 to remove a poison intron, increasing the stability and abundance of PINK1 mRNA and protein. SRSF2P95H-induced mitochondrial dysfunction increased PINK1 expression through this mechanism, which is essential for survival of SRSF2P95H/+ cells. Inhibition of splicing with a glycogen synthase kinase 3 inhibitor promoted retention of the poison intron, impairing mitophagy and activating apoptosis in SRSF2P95H/+ cells. These data reveal a homeostatic mechanism for sensing mitochondrial stress through PINK1 splicing and identify increased mitophagy as a disease marker and a therapeutic vulnerability in SRSF2P95H mutant MDS and AML.

cancer biology↗

bioRxiv2019

Loss of Competence in early development is mediated by loss of chromatin accessibility

As development proceeds, inductive cues are interpreted by competent tissues in a spatially and temporally restricted manner. While key inductive signaling pathways within competent cells are well-described at a molecular level, the mechanisms by which tissues lose responsiveness to inductive signals are not well understood. Localized activation of Wnt signaling before zygotic gene activation in Xenopus laevis leads to dorsal development, but competence to induce dorsal genes in response to Wnts is lost by the late blastula stage. We hypothesize that loss of competence is mediated by changes in histone modifications leading to a loss of chromatin accessibility at the promoters of Wnt target genes. We use ATAC-seq to evaluate genome-wide changes in chromatin accessibility across several developmental stages. Based on overlap with p300 binding, we identify thousands of putative cis-regulatory elements at the gastrula stage, including sites that lose accessibility by the end of gastrulation and are enriched for pluripotency factor binding motifs. Dorsal Wnt target gene promoters are not accessible after the loss of competence in the early gastrula while genes involved in mesoderm and neural crest development maintain accessibility at their promoters. Inhibition of histone deacetylases increases acetylation at the promoters of dorsal Wnt target genes and extends competence for dorsal gene induction by Wnt signaling. Histone deacetylase inhibition, however, is not sufficient to extend competence for mesoderm or neural crest induction. These data suggest that chromatin state regulates the loss of competence to inductive signals in a context-dependent manner.

developmental biology↗