Interactions between HOX Proteins and their Functional Outcomes

Details

Supervisors

Rezsohazy, René ; Gofflot, Françoise

Faculty

Faculté des sciences

Degree label

Master [120] en biochimie et biologie moléculaire et cellulaire, à finalité approfondie

Abstract

In the laboratories of Profs Gofflot and Rezsohazy (LIBST, UCLouvain), it has been observed that some HOX proteins could interact, like HOXA1 with HOXD3 and HOXA5. In some instances, it has also been observed that HOX proteins like HOXA1 can establish homomeric interactions. Such interaction has also been observed with Scr (the HOX homolog of HOX5) in Drosophila. Also, the homomeric interaction of Scr was shown to be required for the good patterning of the fly embryo. Besides these preliminary and fragmentary information, to our knowledge, HOX-HOX interactions have been poorly documented in the literature, and more specifically, the functional consequences for these HOX-HOX interactions remain unexplored. Considering that most HOX genes show overlapping expression patterns in the embryo, HOX-HOX interactions might take place in distinct embryonic territories and cell types. Through this exploratory master thesis, we attempted to answer at least two main questions. First, we wanted to know which HOX protein could interact with another. Second, we wanted to explore if such interactions could influence HOX transcriptional activity. This would therefore contribute to a better understanding of regulatory mechanisms acting on HOX protein activities. To answer the first question, we carried out co-precipitation assays and we focused on HOXA1, HOXA5 and HOXC9 to test whether these proteins could interact with other HOX representatives. To answer the second question, we performed transcriptional activity reporter assays in vitro combining the same HOX proteins to characterize their possible mutual functional influences. Overall, we observed that most HOX pairs tested showed molecular interactions. In addition, our transcriptional activity reporter assays indicated a functional interaction between distinct HOX proteins on the EphA2 enhancer in vitro. Then, while exploring the molecular and the functional interactions focused on HOXA1 -A5 and -C9, we identified that HOXC11 and HOXD11 abrogated the transcriptional activity of other HOX proteins. Next, HOXD13 showed a different functional inhibition towards HOXA5 and HOXC9. Our data therefore suggest that the functional outcomes observed between the different HOX proteins might result from distinct modes of activity interference. Finally, it has been reported that HOX-HOX interactions involving the Drosophila protein Scr relied on a crucial glutamic acid residue within the conserved DNA-binding domain of HOX proteins. We therefore generated mutant HOXA1 and HOXA5 proteins to address the importance of this conserved residue in mammalian for HOX-HOX interactions. We showed that the mutants did not lose their ability to dimerize with their wild type counterpart. However, their transcriptional activity was abrogated. More strikingly, when we co-expressed corresponding mutant and wild type proteins, we observed either a dominant negative (HOXA1 mutants on its wild type) or a positive effect (HOXA5 wild type on the mutants) of their transcriptional activity on the EphA2 enhancer. Taken together these results offer new insights about so far unexplored HOX-HOX molecular and functional interactions.