minibrain

Regulation

The DYRKs (dual specificity tyrosine phosphorylation-regulated kinases) are a conserved family of protein kinases that autophosphorylate a tyrosine residue in their activation loop by an intra-molecular mechanism and phosphorylate exogenous substrates on serine/threonine residues. Little is known about the identity of true substrates for DYRK family members and their binding partners. To address this question, full-length dDYRK2 (Drosophila DYRK2) was used as bait in a yeast two-hybrid screen of a Drosophila embryo cDNA library. Of 14 independent dDYRK2 interacting clones identified, three were derived from the chromatin remodelling factor, SNR1 (Snf5-related 1), and three from the essential chromatin component, TRX (trithorax). The association of dDYRK2 with SNR1 and TRX was confirmed by co-immunoprecipitation studies. Deletion analysis showed that the C-terminus of dDYRK2 modulated the interaction with SNR1 and TRX. DYRK family member MNB (Minibrain) was also found to co-precipitate with SNR1 and TRX, associations that did not require the C-terminus of the molecule. dDYRK2 and MNB were also found to phosphorylate SNR1 at Thr102 in vitro and in vivo. This phosphorylation required the highly conserved DH-box (DYRK homology box) of dDYRK2, whereas the DH-box was not essential for phosphorylation by MNB. This is the first instance of phosphorylation of SNR1 or any of its homologues and implicates the DYRK family of kinases with a role in chromatin remodelling (Kinstrie, 2006. Full text of article).

Riquiqui and Minibrain are regulators of the Hippo pathway downstream of Dachsous

The atypical cadherins Fat (Ft) and Dachsous (Ds) control tissue growth through the Salvador-Warts-Hippo (SWH) pathway, and also regulate planar cell polarity and morphogenesis. Ft and Ds engage in reciprocal signalling as both proteins can serve as receptor and ligand for each other. The intracellular domains (ICDs) of Ft and Ds regulate the activity of the key SWH pathway transcriptional co-activator protein Yorkie (Yki). Signalling from the FtICD is well characterized and controls tissue growth by regulating the abundance of the Yki-repressive kinase Warts (Wts). This study identified two regulators of the Drosophila melanogaster SWH pathway that function downstream of the DsICD: the WD40 repeat protein Riquiqui (Riq) and the DYRK-family kinase Minibrain (Mnb). Ds physically interacts with Riq, which binds to both Mnb and Wts. Riq and Mnb promote Yki-dependent tissue growth by stimulating phosphorylation-dependent inhibition of Wts. Thus, this study describes a previously unknown branch of the SWH pathway that controls tissue growth downstream of Ds (Degoutin, 2013).

The related cadherins Ft and Ds control tissue growth by regulating SWH pathway activity, and also control PCP and morphogenesis. Intriguingly, Ft and Ds regulate SWH pathway activity by engaging in reciprocal signalling as a ligand-receptor pair. Signalling downstream of Ft is reasonably well defined, but signalling downstream of Ds in growth control has remained uncharacterized until the discovery of a membrane-to-nucleus signalling pathway controlling SWH pathway activity downstream of the DsICD, described in this study. Genetic and biochemical data imply that the WD40 repeat protein Riq complexes with the DsICD and can bind to the Mnb and Wts kinases. Riq promotes Mnb-dependent phosphorylation and inhibition of Wts, and thereby promotes Yki-dependent tissue growth. Further investigation is required to define biologically relevant Wts residues that are phosphorylated by Mnb. This study shows that Mnb phosphorylates Wts on several residues in its amino-terminal third, although it is formally possible that other regions of Wts are also phosphorylated by Mnb (Degoutin, 2013).

Therefore, Ds-Ft ligation induces two seemingly opposing growth-regulatory events: Ds activates Ft, which represses Yki by modulating Dachs whereas Ft signals through Ds, Riq and Mnb to activate Yki. At first glance it seems counter-intuitive that Ft-Ds binding would both promote, and repress Yki-dependent tissue growth but raises several interesting possibilities. One option is that the timing of signalling from both the DsICD and the FtICD is different and varies throughout the cell cycle. For example, DsICD might deliver a pulse of Yki activity to induce transcriptional events associated with tissue growth. Subsequently, to ensure that Yki activity does not perdure and cause tissue overgrowth, it could be repressed by signalling from FtICD. Alternatively, DsICD or FtICD signalling might predominate over the other in different regions of imaginal discs or at different stages of development, to regulate Yki. Such regulation could occur through several mechanisms; 1) it could possibly stem from polarized activity of Ft and Ds that occurs in cells of growing imaginal discs in response to graded expression of Ds and Fj, 2) it could occur if the influence of signalling downstream of FtICD or DsICD on Wts activity was quantitatively different, 3) it could result from non-uniform activity of additional proteins that mediate Ft and Ds signalling. Alternatively, repression of Yki by the FtICD, and activation by the DsICD, could quantitatively oppose each other and serve to set a fine threshold of Yki activity that is highly sensitive to regulation by other branches of the SWH pathway such as the Kibra-Ex-Merlin complex, the Hpo activating kinase Tao-1 or apicobasal polarity proteins. In future studies it will be important to define the spatiotemporal activity profile of FtICD and DsICD signalling and the relative influence of the Ds and Ft branches of the SWH pathway on tissue growth (Degoutin, 2013).

Given that Ft and Ds also engage in bi-directional signalling to control PCP and morphogenesis, it will be important to determine whether Riq and Mnb control these processes downstream of the DsICD. In addition, it will be important to investigate whether the signalling events described in this study are conserved in mammals. Interestingly, a reverse regulatory event to that described in this study, between the human orthologues of Wts (LATS2) and Mnb (DYRK1A), has been reported. LATS2 was shown to phosphorylate DYRK1A and promote senescence of cultured cells, raising the possibility that Wts/LATS1/2 and Mnb/DYRK1A/1B kinases engage in mutual regulatory relationships (Degoutin, 2013).

Finally, given the emergence of the SWH pathway as an important regulator of different human tumours, the present study raises the possibility that in a pathological setting the human orthologues of Riq (DCAF7) and Mnb (DYRK1A and DYRK1B) could function as oncogenes. Cell culture studies have provided conflicting reports on whether DYRK1A and DYRK1B act as oncogenes or tumour suppressor genes However, in vivo studies in both flies and mice, and genetic studies in humans, have described only positive roles for Mnb/DYRK1A/DYRK1B in tissue growth: dyrk1a heterozygous mice exhibit growth retardation and impaired brain development, DYRK1A mutations cause microcephaly and growth retardation in humans, whereas Mnb promotes D. melanogaster tissue growth ). These in vivo studies support the possibility that DYRK1A, DYRK1B and DCAF7 could be oncogenic in human cancers (Degoutin, 2013).

DEVELOPMENTAL BIOLOGY

Protein extracts of embryos and pupae contain consistently more Mnb protein A and C than those of third instar larvae and adults. By contrast, Mnb protein B appears to be expressed most markedly in third instar larvae and pupae. In addition, Mnb protein B is the most prominent of the three in third instar larvae (Tejedor, 1995).

Embryonic

In late embryos, MNB mRNA is expressed in the ventral cord and in the brain, but not in the peripheral nervous system. Also, MNB mRNA is not detected in embryonic neuroblasts (Tejedor, 1995).

Larval

Anti Mnb antibodies stain most prominently the mushroom body neuropil and the opc of the optic lobes. Thus mnb appears to be expressed prominently in larval tissue where neuronal progeny are generated during post-embryonic development. Strikingly, the level of protein is low in adult optic lobes and central brain hemispheres (Tejedor, 1995)

Adult

The level of Mnb protein is low in adult optic lobes and central brain hemespheres but relatively high in retinal pigment cells and in the alpha, beta and gama lobes and peduncle of the mushroom bodies (Tejedor, 1995).

Effects of mutation or deletion

Four alleles of minibrain have been described. The external appearance of mutant flies, including body and sensory organs, is nearly indistinguishable from wild type. The mutants are slightly smaller in size and require about 10% more time for their development; they also have considerable difficulties escaping from their pupal case. The brains of adult mutant flies are greatly reduced in size but shows no gross alterations in neuronal architecture. Major size reductions are seen in the optic lobes (50%-70%), most markedly in the lobula complex and in the central brain (40%-50%). The marked reduction of the lobula complex is probably also the reason for the increased curvature of the medulla in mnb mutants. The central brain hemispheres are reduced mainly in their ventral to dorsal and, respectively, anterior and posterior extensions. Axon bundles that project from the lobula complex to the lateral protocerebrum (optic stalk) are visibly thinner in the mutants. The number of anterior optic tract fibers is reduced by about 70%, and the number of cervical connective fibers is reduced by about 30%. Eyes appear normal (Tejedor, 1995).

Freely walking mutant flies cannot fixate a pattern in an area test. Wild type flies are attracted by a vertical dark stripe surrounded by an illuminated translucent area, while mutant flies have lost this preference. Odor discrimination is poor. Although locomotor activity of freely walking animals is low, optomotor turning behavior of mutant males walking on a styrofoam ball and motion-induced landing responses are normal.

minibrain mutant larvae develop normally into the third instar. Mutations cause an abnormal spacing of neuroblasts in the outer proliferation center (opc) of larval brain, with the implication that mnb opc neuroblasts produce less neuronal progeny than do wild type. As a consequence, the adult mnb brain exhibits a specific and marked size reduction of the optic lobes and central brain hemispheres. The insufficient number of distinct neurons in mnb brains is correlated with specific abnormalities in visual and olfactory behavior, although eye and antennal morphology are normal (Tejedor, 1995).

The influence of mutations in seven neurological genes on the number of fibers in the anterior optic tract (AOT) of Drosophila melanogaster has been investigated. The number of fibers in the AOT can be drastically reduced in single and especially in multiple mutants. However, no evidence for synergistic interactions between the sample of mutations used in any of the genes examined (sine oculis , reduced optic lobes, minibrain, and small optic lobes) was obtained at the level of the AOT. The rolKS222 and so mutations eliminate similar fiber sets in the AOT, which are distinctly different from those eliminated by solKS58 and mnb1 (Hoube, 1992).

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date revised: 22 March 2022