Medea SUMOylation restricts the signaling range of the Dpp morphogen in the Drosophila embryo

Abstract

Morphogens are secreted signaling molecules that form concentration gradients and control cell fate in developing tissues. During development, it is essential that morphogen range is strictly regulated in order for correct cell type specification to occur. One of the best characterized morphogens is Drosophila Decapentaplegic (Dpp), a BMP signaling molecule that patterns the dorsal ectoderm of the embryo by activating the Mad and Medea (Med) transcription factors. We demonstrate that there is a spatial and temporal expansion of the expression patterns of Dpp target genes in SUMO pathway mutant embryos. We identify Med as the primary SUMOylation target in the Dpp pathway, and show that failure to SUMOylate Med leads to the increased Dpp signaling range observed in the SUMO pathway mutant embryos. Med is SUMO modified in the nucleus, and we provide evidence that SUMOylation triggers Med nuclear export. Hence, Med SUMOylation provides a mechanism by which nuclei can continue to monitor the presence of extracellular Dpp signal to activate target gene expression for an appropriate duration. Overall, our results identify an unusual strategy for regulating morphogen range that, rather than impacting on the morphogen itself, targets an intracellular transducer. Med and its vertebrate ortholog Smad4 have been found to constitutively shuttle between the nucleus and cytoplasm in a signal-independent manner (Pierreux et al. 2000;Yao et al. 2008). Smad4 contains both a nuclear localization signal (NLS) and a CRM-1-dependent nuclear export signal (NES), and it has been proposed that the relative strengths of these two signals within a tissue regulate the amount of shuttling. Smad4, which shuttles into the nucleus in the absence of signal, is unable to activate transcription (Pierreux et al. 2000), possibly due to recruitment of a repressive complex harboring the SnoN oncoprotein (Stroschein et al. 1999).The small ubiquitin-like modifier protein SUMO is conjugated to its substrate through the sequential activities of E1, E2, and E3 enzymes. Ubc-9, the essential E2 enzyme, catalyzes the conjugation of SUMO to the target lysine, typically located in a ⌿KxE consensus motif (where ⌿ is a large hydrophobic residue and x is any residue) (Hay 2005). An extended SUMO motif compris-

Informatics approaches to understanding TGFβ pathway regulation

“…Given that both lysines, and their contexts, are conserved between H. sapiens SMAD4 and D. melanogaster Medea, Konikoff et al predicted that both lysines are sumoylated in Medea (Konikoff et al, 2008). In the same month, Miles et al demonstrated that Lys141 and Lys185 in Medea are sumoylated and that sumoylation reduces Medea activity in embryonic dorsal/ventral patterning (Miles et al, 2008), validating this prediction. Note that the lysine numbering used in this review and in Konikoff et al derives from GenBank NP 524610 (Wisotzkey et al, 1998;Das et al, 1998), whereas that employed by Miles et al, who report sumoylation of Lys115 and Lys159, derives from GenBank AF039232 (Hudson et al, 1998).…”

Neuroendocrine regulation of Drosophila metamorphosis requires TGFβ/Activin signaling

“…These phenotypes are strikingly similar to those seen in dSmad2 RNAi larvae. Interestingly, previous studies have demonstrated that Medea can be SUMOlyated in vitro, providing a potential link between SUMOlyation and Activin signaling (Miles et al, 2008). However, we have been unable to detect any SUMOylated forms of Medea or dSmad2 under various signaling conditions in vivo or in S2 cells.…”

The regulation of TGFβ signal transduction

“…Sumoylation of SMAD3 by the protein inhibitor of activated Stat y (PIASy; PIAS4) sumoligase promotes its nuclear export in mammalian cells (Imoto et al, 2008). Sumoylation of Medea, by an as yet unidentified sumoligase, also promotes its nuclear export, providing negative regulation that restricts the competence of early Drosophila embryonic cells to respond to Dpp (Miles et al, 2008). This mechanism resembles the previously established role of mammalian SMAD4 sumoylation by PIAS ligases (reviewed by Lönn et al, 2009).…”