Histone Methyltransferase KMT1A Restrains Entry of Alveolar Rhabdomyosarcoma Cells into a Myogenic Differentiated State

Abstract

Alveolar rhabdomyosarcoma (ARMS) is an aggressive pediatric muscle cancer which arrested during the process of skeletal muscle differentiation. In muscle myoblast cells, ectopic expression of the histone H3 lysine 9 (H3K9) methytransferase KMT1A blocks differentiation by repressing a myogenic gene expression program. In this study, we tested the hypothesis that activation of a KMT1A-mediated program of transcriptional repression prevents ARMS cells from differentiating. We investigated whether KMT1A represses the expression of differentiation-associated genes in ARMS cells thereby blocking muscle differentiation. Our results demonstrate that expression of KMT1A is induced in human ARMS cancer cell lines when cultured under differentiation-permissible conditions. shRNA-mediated knock-down of KMT1A decreased anchorage dependent and independent cell proliferation and tumor xenograft growth, increased expression of differentiation-associated genes and promoted the appearance of a terminally differentiated-like phenotype. Lastly, shRNA-directed KMT1A knockdown restored the impaired transcriptional activity of the myogenic regulator MyoD. Together, our results suggested that high levels of KMT1A in ARMS cells under differentiation conditions impairs MyoD function thereby arresting myogenic differentiation in these tumor cells. Thus, targeting KMT1A may be a novel strategy for the treatment of this disease.

The Polycomb group (PcG) protein EZH2 supports the survival of PAX3-FOXO1 alveolar rhabdomyosarcoma by repressing FBXO32 (Atrogin1/MAFbx)

“…Differences in the response to the same treatment of RMS variants are not unusual and are in agreement with both the different cytogenetic and molecular background of fusion-positive alveolar RMS also related to the expression of PAX-FOXO1 chimeric proteins. 1,2,[61][62][63][64][65] Collectively, our data suggest that in PAX3-FOXO1 alveolar RMS EZH2 supports tumor cell survival at least in part by repressing FBXO32.…”

AKT and PAX3-FKHR cooperation enforces myogenic differentiation blockade in alveolar rhabdomyosarcoma cell

“…[25][26][27][28] Despite activated AKT pathway in ARMS cells, they are defective in terminal myogenic differentiation. 17,40,41 In ARMS, it is well recognized that PAX3-FKHR works by the gain of transcriptional power. 2,50,51 The present study was focused on deciphering the molecular mechanism underlying activated AKT signaling in association with PAX3-FKHR transactivation in the suppression of the myogenic differentiation program in ARMS.…”

“…18,21 Here, we show that mouse ARMS cells reflect the defective terminal myogenic phenotype as revealed by no sign of MyHC expression similar to their human counterpart under differentiation conditions (DM). 17,40,41 In this context, studies have implicated that impaired MyoD transcriptional activity-mediated myogenic gene expression, but not the absence of MyoD, is associated with the failure of ARMS cells to differentiate terminally. 31,42,52 However, the data here indicated that impaired MyoD transactivation function is coupled with decreased levels of MyoD when these ARMS cells are under DM.…”

“…Impaired MyoD transcriptional activity-mediated myogenic gene expression is associated with the failure of ARMS cells to differentiate terminally. 17,40,41 To verify that this is also apparent in ARMS-T and ARMS-325 cells, we evaluated MyoD transactivation ability in these ARMS cells compared with C2C12 myoblasts grown in GM or DM following transduction of lentivirus expressing with a MyoD-responsive luciferase reporter (4RE-Luc). 40 The data showed a decrease in MyoD-mediated reporter gene transcription, which is more in ARMS-T-4RE-Luc than ARMS-325-4RE-Luc cells, but, as anticipated, there was an increase of it in C2-4RE-Luc cells 40 grown in DM (Fig.…”

“…17,40,41 We asked whether such a scenario is shared in mouse ARMS cell lines U20497-T (ARMS-T) and U20325 (ARMS-325), derived from PAX3-FKHR knock-in mouse models of ARMS, which contain inactivated INK4a/ARF or p53, 18 respectively. Immunoblot analysis of extracts of these cells confirmed the expression of both PAX3-FKHR and myogenic regulator MyoD proteins, a typical feature of PAX3-FKHR-expressed ARMS cells, 17,18,[42][43][44] with less PAX3-FKHR and more MyoD in ARMS-325 cells than the ARMS-T cells (Fig.…”