Absence of Bcl11b in murine heart leads to mitochondrial dysfunction and cardiomyocyte death

Abstract

B cell lymphoma 11b (Bcl11b/Ctip2) is a zinc finger transcriptional factor involved in a number of physiological processes, including cardiac hypertrophy. In humans, genome-wide association studies show that a common genetic variation in the 3’ region of Bcl11b gene is associated with higher risk of aortic stiffness and increased rate of cardiovascular disease. In mice, Bcl11b invalidation specifically in cardiomyocytes leads to cardiac malfunction, fibrosis augmentation and cell death. Our specific aims are: To investigate the signalling pathways related to cardiac cell death and to assess the sarcomere structure of cardiomyocytes in the absence of Bcl11b. The experiments were conducted using cardiac specific Bcl11b knock-out mice induced by tamoxifen administration 3 and 7 days after the invalidation, cardiac tissues were collected to perform RNA sequencing and subsequently to analyse related molecular pathways using bioinformatic tools. In parallel, cardiomyocytes were purified and isolated by conducting Langendorff perfusion technique to have a better insight into the impact of Bcl11b. A simplified mathematical model, Fourier transform analysis was used to assess sarcomeric length following Bcl11b invalidation in cardiomyocytes. Based on the RNA sequencing results, Bcl11b invalidation has an impact on the regulation of over 2000 genes. Among them, genes related to cell death and mitochondrial functions are the most affected regarding the enrichment scores. Interestingly, a considerable number of genes fall into the “Cuproptosis” pathway, a newly identified, non-apoptotic cell death mechanism. The oxygen consumption of mitochondria was also measured using the Seahorse assay. Bcl11b deletion leads to the decrease in basal level of mitochondrial respiration with no effect on the maximum respiratory capacity. Finally, adult cardiomyocytes in the mutant groups showed a moderate hypertrophy (15% bigger than the control group). However, Fourier transform analysis exhibited stability in the average sarcomere length of these cells. The obtained data suggest that cardiac specific invalidation of Bcl11b induces mitochondrial dysfunction and possibly cell death through the “Cuproptosis” pathway.