A poly-histidine motif of HOXA1 is involved in regulatory interactions with cysteine-rich proteins.

Homopolymeric amino acid repeats are found in about 24 % of human proteins and are over-represented in transcriptions factors and kinases. Although relatively rare, homopolymeric histidine repeats (polyH) are more significantly found in proteins involved in the regulation of embryonic development. To gain a better understanding of the role of polyH in these proteins, we used a bioinformatic approach to search for shared features in the interactomes of polyH-containing proteins in human. Our analysis revealed that polyH protein interactomes are enriched in cysteine-rich proteins and in proteins containing (a) cysteine repeat(s). Focusing on HOXA1, a HOX transcription factor displaying one long polyH motif, we identified that the polyH motif is required for the HOXA1 interaction with such cysteine-rich proteins. We observed a correlation between the length of the polyH repeat and the strength of the HOXA1 interaction with one Cys-rich protein, MDFI. We also found that metal ion chelators disrupt the HOXA1-MDFI interaction supporting that such metal ions are required for the interaction. Furthermore, we identified three polyH interactors which down-regulate the transcriptional activity of HOXA1. Taken together, our data point towards the involvement of polyH and cysteines in regulatory interactions between proteins, notably transcription factors like HOXA1.

HOXA2 activity regulation by cytoplasmic relocation, protein stabilization and post-translational modification.

HOX proteins are homeodomain transcription factors critically involved in patterning animal embryos and controlling organogenesis. While the functions of HOX proteins and the processes under their control begin to be well documented, the modalities of HOX protein activity regulation remain poorly understood. Here we show that HOXA2 interacts with PPP1CB, a catalytic subunit of the Ser/Thr PP1 phosphatase complex. This interaction co-localizes in the cytoplasm with a previously described HOXA2 interactor, KPC2, which belongs to the KPC E3 ubiquitin ligase complex. We provide evidence that HOXA2, PPP1CB and KPC2 define a molecularly and functionally interacting complex. Collectively, our experiments support that PPP1CB and KPC2 together inhibit the activity of HOXA2 by activating its nuclear export, but favored HOXA2 de-ubiquitination and stabilization thereby establishing a store of HOXA2 in the cytoplasm.

PRDM14, a putative histone methyl-transferase, interacts with and decreases the stability and activity of the HOXA1 transcription factor.

Understanding how the activity of transcription factors like HOX proteins is regulated remains a widely open question. In a recent screen for proteins interacting with HOXA1, we identified a PRDM protein family member, PRDM14, which is known to be transiently co-expressed with HOXA1 in epiblast cells before their specification towards somatic versus germ cell fate. Here, we confirm PRDM14 is an interactor of HOXA1 and we identify the homeodomain of HOXA1 as well as the PR domain and Zinc fingers of PRDM14 to be required for the interaction. An 11-His repeat of HOXA1 previously highlighted to contribute to HOXA1-mediated protein-protein interactions is also involved. At a functional level, we provide evidence that HOXA1 displays an unexpectedly long half-life and demonstrate that PRDM14 can reduce the stability and affect the transcriptional activity of HOXA1.

The O-GlcNAc transferase OGT interacts with and post-translationally modifies the transcription factor HOXA1.

HOXA1 belongs to the HOX family of transcription factors which are key regulators of animal development. Little is known about the molecular pathways controlling HOXA1. Recent data from our group revealed distinct partner proteins interacting with HOXA1. Among them, OGT is an O-linked N-acetylglucosamine (O-GlcNAc) transferase modifying a variety of proteins involved in different cellular processes including transcription. Here, we confirm OGT as a HOXA1 interactor, we characterise which domains of HOXA1 and OGT are required for the interaction, and we provide evidence that OGT post-translationally modifies HOXA1. Mass spectrometry experiments indeed reveal that HOXA1 can be phosphorylated on the AGGTVGSPQYIHHSY peptide and that upon OGT expression, the phosphate adduct is replaced by an O-GlcNAc group.