Identification of ZBTB24 protein domains and motifs for heterochromatin localization and transcriptional activation

Abstract

Immunodeficiency, centromeric instability, facial anomalies (ICF) syndrome is a rare autosomal recessive disorder caused by mutations in either DNMT3B, ZBTB24, CDCA7, HELLS or an unknown gene(s). Among the known causative genes, ZBTB24 encodes a member of the BTB‐zinc finger (ZF) transcription factor family. The protein possesses a BTB domain, an AT‐hook and eight C2H2 ZF motifs. All ZBTB24 mutations reported in ICF patients are predicted to disrupt at least one ZF motif. Here, we show that both AT‐hook and distinct ZF motifs, particularly the 6th motif, of human and mouse ZBTB24 proteins are important for their heterochromatin localization. On the other hand, the 6th and 7th ZF motifs, and not the AT‐hook or the BTB domain, of the human and mouse proteins are essential for transcriptional activation of CDCA7, another ICF causative gene and a known target of ZBTB24. By deletion analysis of the human CDCA7 promoter, we show that two motifs for ZBTB24 binding are important for transcriptional activation of this gene. These results reveal the evolutionarily conserved domains and motifs important for the biological function of ZBTB24, which provides a basis for understanding the molecular mechanisms underlying the pathogenesis of ICF syndrome.

ZBTB24 is a conserved multifaceted transcription factor at genes and centromeres that governs the DNA methylation state and expression of satellite repeats

“…S1A and S2A-S2D ). Of note, the ZBTB24 protein sequence is highly conserved between mouse and human ranging from 76.9 to 96.9 % of homology for the structural domains (26).…”

“…However, as the AT-hook domain alone has a low affinity for DNA, the affinity of ZBTB24 for centromeric satellite sequences would be potentiated by the joint action of both AT-hook and ZnF motifs (57). In line with this, the presence of both motifs has been shown to be important for the localization of ZBTB24 to (peri)centromeric heterochromatin (26). Considering the growing body of evidence that points to a role of DNAme in centromeric integrity (4) and the fundamental role of centromeres in ensuring faithful transmission of the genetic information (58), ZBTB24 emerges as a guardian of centromeric integrity and hence, of genomic stability.…”

“…Interestingly, in murine NIH3T3 cells, exogenous ZBTB24 protein localizes to chromocenters, i.e. subnuclear clusters of (peri)centromeres from several chromosomes, suggesting that ZBTB24 would also play an important role at (peri)centromeric heterochromatin (25, 26). As shown in zebrafish, ZBTB24 function would be essential for the methylated status of pericentromeric satellite DNA repeats and their transcriptional repression (27).…”

Zbtb24 binding protects promoter activity by antagonizing DNA methylation in mESCs

“…Interestingly, a very recent study showed that both the AT-hook and the ZF domain of ZBTB24 are required for heterochromatin localization of the protein but how ZBTB24 gets recruited to these sites remains unclear (Aktar et al 2019). It has also been shown that Zbtb24 associates with pericentromeric heterochromatin in NIH3T3 cells independent of DNA methylation (Nitta et al 2013).…”

“…A combined function of ZBTB24 and DNMT3B has been suggested for mediating some gene body methylation in HCT116 cells (Thompson et al 2018), but the mechanisms by which ZBTB24 influences DNA methylation at other genomic regions remain unclear. ZBTB24 also functions as a transcriptional activator/repressor, and with its C2H2 zinc finger domain, ZBTB24 can bind to DNA in a sequence-specific manner (Thompson et al 2018;Aktar et al 2019;Ren et al 2019).…”