BMP signalling: development, stem cell maintenance and cancer
The Bone Morphogenic Proteins (BMPs) represent a major, highly conserved family of growth factor signalling molecules. The BMP signalling pathway is a key regulator of a diverse array of processes, including the patterning of many different organs and tissues during embryonic development. As a consequence, misregulation of BMP signalling is associated with a spectrum of human diseases, including different cancer types, vascular disorders, kidney diseases and skeletal defects. The research in my lab uses the genetically tractable Drosophila model organism to investigate the regulation of BMP signaling. In Drosophila the major BMP signalling molecule is called Dpp, and Dpp signalling is important during numerous stages of development. The primary research interest of my lab is to understand how Dpp signalling is regulated in the ovary and early embryo. In the ovary Dpp is a critical signal required to maintain germline stem cells, whereas in the embryo a gradient of Dpp activity leads to the specification of different cell types. The lab also has an interest in the regulation of translation during oogenesis.
Research projects are as follows:
Extracellular regulation of Dpp signalling
Collagen IV and Integrins: We have found that type IV collagens, encoded by the vkg and Dcg1 genes in Drosophila, bind Dpp and augment its signaling in the early embryo (Wang et al, 2008, Nature 454, 72). More recently, we have mapped additional protein interactions allowing us to formulate a molecular model of Dpp shuttling complex assembly on collagen IV. We provide in vivo support for the key finding relating to this model, that Dpp is more diffusible in the embryo when it cannot bind collagen IV (Sawala et al, 2012, PNAS 109,11122). We have also extended our analysis by testing the role of integrins. Our data are consistent with integrin signalling acting downstream of collagen IV to enhance the Dpp pathway. Current work is aimed at understanding the molecular mechanism by which integrins amplify the Dpp pathway.
Tolloid protease: Dpp gradient formation requires the assembly of a shuttling complex that promotes movement of the active Dpp/Scw heterodimer. This Dpp/Scw-Sog-Tsg complex is inhibitory with respect to Dpp signaling, until Dpp is liberated from the complex by Tolloid cleavage of the Sog protein. In collaboration with Clair Baldock we are undertaking a structure-function analysis of Drosophila Tolloid. We are using biophysical data to make predictions about Tolloid activity that are then tested in the embryo.
Transcriptional regulation in response to the Dpp signal
Mad, Brk transcriptional network: We are using ChIP-seq and other whole-genome approaches to determine the Dpp responsive transcriptional programme at different time points during embryogenesis. We are using the data to identify new motifs in Dpp-responsive enhancers, determine how features of the enhancers correlate with expression pattern, and gain mechanistic insight into Dpp target gene activation.
Regulation of Smad and Brk activity: We have used tissue culture RNAi screens to identify new regulators of Mad activation and Brk repression. Current work is aimed at validating potential targets in the embryo. One aim is to integrate these data sets with the Mad and Brk ChIP-seq data in order to identify tiers of regulation that shape Dpp-responsive transcription. We are also interested in how the SUMO pathway impacts on Dpp signalling in the embryo, having shown that SUMO modification of the Med transcription factor negatively regulates its activity (Miles et al, 2008, Genes Dev 22, 2578).
Post-transcriptional regulation of stem cell fate
Translational control and miRNAs: Maintenance of ovarian germline stem cells (GSCs) requires the Pumilio-Nanos (Pum-Nos) translational repressors. Upon GSC division, one daughter remains a GSC whereas the other differentiates into a cystoblast (CB). The Pum-Nos complex is active in GSCs, but in CBs the translation of Nanos is repressed, leading to a loss of Pum-Nos repression. We have shown that Pum-Nos repress translation of the brat mRNA, which encodes a translational repressor, in GSCs (Harris et al, 2011, Dev Cell20, 72). This repression is alleviated in CBs, allowing formation of a Pum-Brat translation repression complex. One of the mRNAs repressed by Pum-Brat is the Mad mRNA, which extinguishes the CB’s ability to respond to the self-renewal Dpp signal. Current work is aimed at identifying additional targets of Pum-Nos repression in GSCs, as well as more detailed insight into their repressive mechanism. Given the conserved role of Puf and Nos proteins in other stem cell systems, it is anticipated that our data will provide a framework for understanding how translation regulation impacts upon stem cell fate choices in other organisms, including humans, where there is vast therapeutic potential to be harnessed through the precise manipulation of stem cell fate. The miRNA pathway is essential for GSC maintenance, although the role of individual miRNAs is poorly understood. We are interested in identifying the mRNA targets of specific miRNAs in GSCs and CBs and understanding how repression of these mRNAs impacts upon the GSC/CB cell fate choice.