Apoptotic cell death removes unwanted cells and is regulated by interactions between pro-survival and pro-apoptotic members of the BCL-2 protein family. The regulation of apoptosis is thought to be crucial for normal embryonic development. Accordingly, complete loss of pro-survival MCL-1 or BCL-XL (BCL2L1) causes embryonic lethality. However, it is not known whether minor reductions in pro-survival proteins could cause developmental abnormalities. We explored the rate-limiting roles of MCL-1 and BCL-XL in development and show that combined loss of single alleles of Mcl-1 and Bcl-x causes neonatal lethality. Mcl-1;Bcl-x mice display craniofacial anomalies, but additional loss of a single allele of pro-apoptotic Bim (Bcl2l11) restores normal development. These findings demonstrate that the control of cell survival during embryogenesis is finely balanced and suggest that some human craniofacial defects, for which causes are currently unknown, may be due to subtle imbalances between pro-survival and pro-apoptotic BCL-2 family members.

HBO1 (MYST2/KAT7) is essential for histone 3 lysine 14 acetylation (H3K14ac) but is dispensable for H4 acetylation and DNA replication in mouse tissues. In contrast, previous studies using small interfering RNA (siRNA) knockdown in human cell lines have suggested that HBO1 is essential for DNA replication. To determine if HBO1 has distinctly different roles in immortalized human cell lines and normal mouse cells, we performed siRNA knockdown of HBO1. In addition, we used CRISPR/Cas9 to generate 293T, MCF7, and HeLa cell lines lacking HBO1. Using both techniques, we show that HBO1 is essential for all H3K14ac in human cells and is unlikely to have a direct effect on H4 acetylation and only has minor effects on cell proliferation. Surprisingly, the loss of HBO1 and H3K14ac in HeLa cells led to the secondary loss of almost all H4 acetylation after 4 weeks. Thus, HBO1 is dispensable for DNA replication and cell proliferation in immortalized human cells. However, while cell proliferation proceeded without HBO1 and H3K14ac, HBO1 gene deletion led to profound changes in cell adhesion, particularly in 293T cells. Consistent with this phenotype, the loss of HBO1 in both 293T and HeLa principally affected genes mediating cell adhesion, with comparatively minor effects on other cellular processes.

Histone acetylation is essential for initiating and maintaining a permissive chromatin conformation and gene transcription. Dysregulation of histone acetylation can contribute to tumorigenesis and metastasis. Using inducible cre-recombinase and CRISPR/Cas9-mediated deletion, we investigated the roles of the histone lysine acetyltransferase TIP60 (KAT5/HTATIP) in human cells, mouse cells, and mouse embryos. We found that loss of TIP60 caused complete cell growth arrest. In the absence of TIP60, chromosomes failed to align in a metaphase plate during mitosis. In some TIP60 deleted cells, endoreplication occurred instead. In contrast, cell survival was not affected. Remarkably, the cell growth arrest caused by loss of TIP60 was independent of the tumor suppressors p53, INK4A and ARF. TIP60 was found to be essential for the acetylation of H2AZ, specifically at lysine 7. The mRNA levels of 6236 human and 8238 mouse genes, including many metabolism genes, were dependent on TIP60. Among the top 50 differentially expressed genes, over 90% were downregulated in cells lacking TIP60, supporting a role for TIP60 as a key co-activator of transcription. We propose a primary role of TIP60 in H2AZ lysine 7 acetylation and transcriptional activation, and that this fundamental role is essential for cell proliferation. Growth arrest independent of major tumor suppressors suggests TIP60 as a potential anti-cancer drug target.

The Börjeson–Forssman–Lehmann syndrome (BFLS) is an intellectual disability and endocrine disorder caused by plant homeodomain finger 6 (PHF6) mutations. BFLS patients present with short stature. We report a mouse model of BFLS, in which deletion of Phf6 causes a proportional reduction in body size compared to control mice. Growth hormone (GH) levels were reduced in the absence of PHF6. Phf6−/Y animals displayed a reduction in the expression of the genes encoding GH releasing hormone (GHRH) in the brain, GH in the pituitary gland and insulin-like growth factor-1 (IGF-1) in the liver. Phf6 deletion specifically in the nervous system caused a proportional growth defect, indicating neuroendocrine contribution to the phenotype. Loss of suppressor of cytokine signaling 2 (SOCS2), a negative regulator of growth hormone signaling partially rescued body size, supporting a reversible deficiency in GH signaling. These results demonstrate that PHF6 regulates the GHRH/GH/IGF-1 axis.

No abstract