Research Article: Promotion of adipogenesis by JMJD6 requires the AT hook-like domain and is independent of its catalytic function

Date Published: August 20, 2019

Publisher: Public Library of Science

Author(s): Pablo Reyes-Gutierrez, Jake W. Carrasquillo-Rodríguez, Anthony N. Imbalzano, Fenfei Leng.


JMJD6 is a member of the Jumonji C domain containing enzymes that demethylate and/or hydroxylate substrate proteins. It is a multi-functional protein that has been implicated in disparate aspects of transcriptional and post-transcriptional control of gene expression, including but not limited to enhancer and promoter binding, release of paused RNA polymerase II, control of splicing, and interaction with the translation machinery. JMJD6 contributes to multiple aspects of animal development, including adipogenesis modeled in culture. We mutated proposed or characterized domains in the JMJD6 protein to better understand the requirement for JMJD6 in adipogenic differentiation. Mutation of JMJD6 amino acids that mediate binding of iron and 2-oxogluterate, which are required cofactors for enzymatic activity, had no impact on JMJD6 function, showing that catalytic activity is not required for JMJD6 contributions to adipogenic differentiation. In addition, we documented the formation of JMJD6 oligomers and showed that catalytic activity is not required for oligomerization, as has been reported previously. We also observed no effect of mutations in the sumoylation site and in the poly-serine stretch. In contrast, mutation of the AT hook-like structure, which mediates interaction with DNA and/or RNA, compromised JMJD6 function by blocking its ability to interact with chromatin at genes that express regulators of adipogenesis. The ability of JMJD6 to interact with nucleic acids may be a critical requirement for its function in adipogenic differentiation. The requirement for the AT hook-like domain and the lack of requirement for catalytic activity giving rise to the idea that co-activation of transcription by JMJD6 may be functioning as a scaffold protein that supports the interactions of other critical regulators.

Partial Text

The Jumonji (Jmj) family of proteins encodes evolutionarily conserved oxygenases dependent on ferrous iron (Fe2+) and 2-oxogluterate to hydroxylate metabolites, proteins and nucleic acids [1, 2]. The conserved JmjC domain is structurally related to the cupin domains found in archaea and other kingdoms that possess active sites containing a metal ion within a histidine cluster [3]. JmjC domains form a double-stranded β-helical fold in which eight β-strands form two, four-stranded antiparallel β-sheets [4]. Distinctions between different JmjC families are generally defined by the structural elements that surround the conserved JmjC domain and the presence of other protein domains, many of which are interaction surfaces for chromatin or chromatin-bound proteins. The JmjC proteins themselves are overwhelmingly described as factors that promote the regulation of transcription and/or chromatin [5].

We previously examined the role of JMJD6 and determined it promoted adipogenesis in culture through both transcriptional and post-transcriptional mechanisms [11]. In that study, we showed that shRNA-mediated knockdown of Jmjd6 inhibited adipogenesis and that ectopic expression of JMJD6 could rescue the differentiation deficiency. We also rescued the shRNA-induced deficiency in differentiation by expressing a JMJD6 protein mutated at amino acids 187 and 189, which are critical for iron binding [11]. Others have shown that mutations in one or both of these amino acids significantly compromise or abolish JMJD6 enzymatic activity [10, 24, 39, 40, 44, 46]. Our results suggest that the catalytic activity of JMJD6 is not necessary for its role in adipogenesis and raise the question of which JMJD6 domains are required for this function.