Academic journal article Genetics

Alternative Splicing Modulates Ubx Protein Function in Drosophila Melanogaster

Academic journal article Genetics

Alternative Splicing Modulates Ubx Protein Function in Drosophila Melanogaster

Article excerpt

ABSTRACT

The Drosophila Hox gene Ultrabithorax (Ubx) produces a family of protein isoforms through alternative splicing. Isoforms differ from one another by the presence of optional segments-encoded by individual exons-that modify the distance between the homeodomain and a cofactor-interaction module termed the "YPWM" motif. To investigate the functional implications of Ubx alternative splicing, here we analyze the in vivo effects of the individual Ubx isoforms on the activation of a natural Ubx molecular target, the decapentaplegic (dpp) gene, within the embryonic mesoderm. These experiments show that the Ubx isoforms differ in their abilities to activate dpp in mesodermal tissues during embryogenesis. Furthermore, using a Ubx mutant that reduces the full Ubx protein repertoire to just one single isoform, we obtain specific anomalies affecting the patterning of anterior abdominal muscles, demonstrating that Ubx isoforms are not functionally interchangeable during embryonic mesoderm development. Finally, a series of experiments in vitro reveals that Ubx isoforms also vary in their capacity to bind DNA in presence of the cofactor Extradenticle (Exd). Altogether, our results indicate that the structural changes produced by alternative splicing have functional implications for Ubx protein function in vivo and in vitro. Since other Hox genes also produce splicing isoforms affecting similar protein domains, we suggest that alternative splicing may represent an underestimated regulatory system modulating Hox gene specificity during fly development.

HOX genes are key genetic elements for the development of most animals. They encode a family of transcriptional regulators that trigger different developmental programs at specific coordinates along the antero-posterior body axis of animals (McGinnis and Krumlauf 1992; Alonso 2002; Pearson et al. 2005). In addition to their crucial developmental roles, the biological relevance of Hox genes is further emphasized by the fact that changes in the expression and function of Hox genes are associated with the emergence of new structures during animal evolution, suggesting that genetic variation affecting Hox gene activity may have been causal to the generation of morphological diversity (Lewis 1978; Holland and Garcia-Fernandez 1996; Carroll et al. 2001; Galant and Carroll 2002; Hughes and Kaufman 2002; Ronshaugen et al. 2002; Pearson et al. 2005).

In spite of their generality and importance for animal development and evolution, many aspects of Hox gene function remain unclear. One of them concerns the mechanisms underlying their specificity of action within developmental units, such as the individual segments in the insects. The problem could be stated as follows: given that from very early in development all cells within each segment are allocated a unique Hox "code" (Lewis 1978), all themorphogenetic complexity internal to the segment must be derived from a relatively simple Hox input, leading to the question of how Hox activities could be subspecified at the cellular level during the progressive development of segments. Combinatorial control by different Hox genes acting on overlapping target tissues may be one of the mechanisms used to increase regulatory complexity (Struhl 1982); however, in many cases Hox gene products were shown to compete with each other to take control of a developing segment (Gonzalez-Reyes and Morata 1990; Gonzalez-Reyes et al. 1990; Lamka et al. 1992; Akam 1998), arguing for the existence of other mechanisms for Hox specificity. Cell-specific "cofactors" able to modulate Hox transcriptional functions in individual cells could also provide an important contribution to the specificity of Hox genes (Mann 1995; Mann and Chan 1996). Nonetheless, to date, only a very limited number of Hox cofactors have been identified, making it difficult to explain how complex morphogenetic pathways, which rely on the orchestrated behavior of thousands of cells, could be obtained from such a simple Hox/cofactor molecular code. …

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