In developing from an embryo to an adult, higher organisms accumulate mass at a rapid rate. Once each organ or organ primordium has reached the requisite size, cell proliferation and growth ceases in most of the tissue and will remain confined to specialised cells (such as stem cells) to maintain homeostasis throughout adult life. In order to achieve consistent organ and body size in individuals of the same species, tight control must be applied to cell growth and cell number at the end of development. Since the genes that restrict organ size are likely to be targets of tumour-promoting mutations, the study of these mechanisms is relevant to both developmental and cancer biology.
The Hippo (Hpo) pathway comprises the kinases Hpo and Warts, the adaptors Salvador and Mats, the cytoskeletal proteins Expanded and Merlin, the atypical cadherin Fat and the transcriptional co-factor Yorkie (Harvey et al, Nat Rev Cancer 2007; 7: 182-191). This pathway has been shown to restrict tissue size through the control of cell division and apoptosis during development in Drosophila. The immediate aim of the lab is to identify new members of the Hpo pathway and to understand how the pathway functions at the biochemical level. The lab uses a combination of genetics, cell biology and biochemistry in order to understand how the Hpo pathway fits in the 'big picture' of overall size regulation in development and cancer.
The Hippo pathway and cell polarity
In addition to their well-characterised overproliferation phenotype, epithelial cells mutant for the kinases Hippo and Warts present a hypertrophy of the apical domain. We have examined the molecular basis of this apical hypertrophy and its impact on cell proliferation. In the wing imaginal disc epithelium, we observe increased staining for the apical polarity complexes, such as DaPKC/Par3/Par6 and Crumbs/Stardust when Hippo activity is compromised, while baso-lateral markers are not affected. The cell surface localisation of the Notch receptor is also increased in mutant clones, opening the possibility that aberrant receptor signalling may participate in overgrowth of hpo-deficient tissue. Interestingly however, while the polarity determinant Crumbs is required for the accumulation of apical proteins, this does not appear to significantly contribute to the overproliferation defect elicited by loss of Hippo signalling. Therefore, Hippo signalling controls polarity and growth via distinct mechanisms.
Disruption of epithelial architecture and loss of cell polarity is a hallmark of cancer. In breast or colon cancer, loss of polarised architecture is usually the first sign of transformation (Wodarz et al, Nat Cell Biol 2007; 9: 1016-1024). Can loss of polarity therefore lead to cancer? Recent work on the neoplastic tumour suppressor genes (nTSGs) in Drosophila has put the link between polarity abnormalities and tumour formation into sharp focus (Hariharan et al, Annu Rev Genet 2006; 40: 335-361). nTSGs such as the basal determinants scrib, dlg and lgl present a strong expansion of their apical domain, with ectopic localisation of apical proteins on lateral membrane and ectopic formation of adherens junctions. Those cells also have a massive overproliferation defect: they do not cycle faster than wild-type cells but they do not respond to arrest cues and carry on dividing. However, several studies suggest that, as is the case for Hpo pathway mutants, the proliferative and polarity defects are separable. Thus, there may be more similarities between neoplastic TSGs and hyperplastic TSGs such as Hpo pathway members than previously thought.
Drosophila MFAP1 is required for pre-mRNA processing and G2/M progression
In a non-Hippo related project, we have characterised a novel component of the spliceosome and its function in cell cycle regulation. The mammalian spliceosome has mainly been studied using proteomics. The isolation and comparison of different splicing intermediates has revealed the dynamic association of more than 200 splicing factors with the spliceosome, relatively few of which have been studied in detail. Here, we report the characterisation of the Drosophila homologue of Microfibril-Associated Protein 1 (dMFAP1), a previously uncharacterised protein found in some human spliceosomal fractions (Jurica et al, Mol Cell 2003; 12: 5-14). We showed that dMFAP1 binds directly to the Drosophila homologue of Prp38p (dPrp38), a tri-snRNP component, and is required for pre-mRNA processing (Andersen et al, J Biol Chem 2008; 283: 31256-31267). dMFAP1, like dPrp38, is essential for viability, and our in vivo data show that cells with reduced levels of dMFAP1 or dPrp38 proliferate more slowly than normal cells and undergo apoptosis. Consistent with this, dsRNA-mediated depletion of dPrp38 or dMFAP1 causes cells to arrest in G2/M, and this is paralleled by a reduction in mRNA levels of the mitotic phosphatase string/cdc25. Interestingly dsRNA-mediated depletion of a wide range of core splicing factors elicits a similar phenotype, suggesting that the observed G2/M arrest might be a general consequence of interfering with spliceosome function.
Figure 1. Inactivation of Hpo pathway members induces apical protein accumulation. Transverse sections of a wing imaginal disc. Apical is to the top. Cells mutant for wts (negative for GFP, green in C) present an increase in the apical marker DaPKC (A, and blue in C) but not the lateral marker Dlg (B, and red in C).
For a list of refereed research papers, see Publications (in navigation on left).
