CHARACTERIZATION OF PROTEIN-AND DNA-INTERACTING DOMAINS IN HUMAN DNA (CYTOSINE-5)-METHYLTRANSFERASE

Ph.DDOCTOR OF PHILOSOPH

The multiple interactions of RUNX with the hippo–YAP pathway

10.3390/cells10112925Cells1011292

The RUNX1 Enhancer Element eR1: A Versatile Marker for Adult Stem Cells

10.14348/molcells.2019.0250MOLECULES AND CELLS432121-12

Protocol for identification and validation of IGF2BP1 target genes in pluripotent human embryonic carcinoma cells

Summary: We present a detailed protocol to identify and validate IGF2BP1 target genes in pluripotent human embryonic carcinoma cells (NTERA-2). We first identify the target genes through RNA-immunoprecipitation (RIP) sequencing. We then validate the identified targets through the use of RIP-qPCR assays, determine the m6A status of target genes by m6A-IP, and perform functional validation by quantifying changes in mRNA or protein expression levels upon knockdown of IGF2BP1 or methyltransferases in NTERA-2.For complete details on the use and execution of this protocol, please refer to Myint et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics

RUNX3 in Stem Cell and Cancer Biology

10.3390/cells12030408CELLS12

Aurora kinase-induced phosphorylation excludes transcription factor RUNX from the chromatin to facilitate proper mitotic progression

The Runt-related transcription factors (RUNX) are master regulators of development and major players in tumorigenesis. Interestingly, unlike most transcription factors, RUNX proteins are detected on the mitotic chromatin and apparatus, suggesting that they are functionally active in mitosis. Here, we identify key sites of RUNX phosphorylation in mitosis. We show that the phosphorylation of threonine 173 (T173) residue within the Runt domain of RUNX3 disrupts RUNX DNA binding activity during mitotic entry to facilitate the recruitment of RUNX proteins to mitotic structures. Moreover, knockdown of RUNX3 delays mitotic entry. RUNX3 phosphorylation is therefore a regulatory mechanism for mitotic entry. Cancer-associated mutations of RUNX3 T173 and its equivalent in RUNX1 further corroborate the role of RUNX phosphorylation in regulating proper mitotic progression and genomic integrity.NRF (Natl Research Foundation, S’pore)MOE (Min. of Education, S’pore)Accepted versio

RUNX Poly(ADP-Ribosyl)ation and BLM Interaction Facilitate the Fanconi Anemia Pathway of DNA Repair

Summary: The Fanconi anemia (FA) pathway is a pivotal genome maintenance network that orchestrates the repair of DNA interstrand crosslinks (ICLs). The tumor suppressors RUNX1 and RUNX3 were shown to regulate the FA pathway independent of their canonical transcription activities, by controlling the DNA damage-dependent chromatin association of FANCD2. Here, in further biochemical characterization, we demonstrate that RUNX3 is modified by PARP-dependent poly(ADP-ribosyl)ation (PARylation), which in turn allows RUNX binding to DNA repair structures lacking transcription-related RUNX consensus motifs. SILAC-based mass spectrometric analysis revealed significant association of RUNX3 with core DNA repair complexes, including PARP1, even in unstressed cells. After DNA damage, the increased interaction between RUNX3 and BLM facilitates efficient FANCD2 chromatin localization. RUNX-Walker motif mutations from breast cancers are impaired for DNA damage-inducible PARylation, unveiling a potential mechanism for FA pathway inactivation in cancers. Our results reinforce the emerging paradigm that RUNX proteins are tumor suppressors with genome gatekeeper function. : Tay et al. demonstrate that the tumor suppressor genes RUNX1 and RUNX3 have an important regulatory role in the genome maintenance pathway controlled by FANCD2. DNA damage induces PARP-dependent PARylation of RUNX proteins, leading to their interaction with BLM to control the loading of FANCD2 on DNA damage sites. Keywords: RUNX1, RUNX3, Fanconi anemia, interstrand crosslink repair, PARP1, poly(ADP-ribosyl)ation, BLM, SILAC, DNA repair, FANCD

RUNX3 functions as an oncogene in ovarian cancer

10.1016/j.ygyno.2011.04.044Gynecologic Oncology1222410-417GYNO