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Control of lysosomal biogenesis and Notch-dependent tissue patterning by components of the TFEB-V-ATPase axis in Drosophila melanogaster.

Tognon E, Kobia F, Busi I, Fumagalli A, De Masi F, Vaccari T - Autophagy (2016)

Bottom Line: In vertebrates, TFEB (transcription factor EB) and MITF (microphthalmia-associated transcription factor) family of basic Helix-Loop-Helix (bHLH) transcription factors regulates both lysosomal function and organ development.Similar to our findings in Drosophila, in human breast epithelial cells we observe that impairment of the Vha16-1 human ortholog ATP6V0C changes the size and function of the endolysosomal compartment and that depletion of TFEB reduces ligand-independent N signaling activity.Our data suggest that lysosomal-associated functions regulated by the TFEB-V-ATPase axis might play a conserved role in shaping cell fate.

View Article: PubMed Central - PubMed

Affiliation: a IFOM - FIRC Institute of Molecular Oncology , Milan , Italy.

ABSTRACT
In vertebrates, TFEB (transcription factor EB) and MITF (microphthalmia-associated transcription factor) family of basic Helix-Loop-Helix (bHLH) transcription factors regulates both lysosomal function and organ development. However, it is not clear whether these 2 processes are interconnected. Here, we show that Mitf, the single TFEB and MITF ortholog in Drosophila, controls expression of vacuolar-type H(+)-ATPase pump (V-ATPase) subunits. Remarkably, we also find that expression of Vha16-1 and Vha13, encoding 2 key components of V-ATPase, is patterned in the wing imaginal disc. In particular, Vha16-1 expression follows differentiation of proneural regions of the disc. These regions, which will form sensory organs in the adult, appear to possess a distinctive endolysosomal compartment and Notch (N) localization. Modulation of Mitf activity in the disc in vivo alters endolysosomal function and disrupts proneural patterning. Similar to our findings in Drosophila, in human breast epithelial cells we observe that impairment of the Vha16-1 human ortholog ATP6V0C changes the size and function of the endolysosomal compartment and that depletion of TFEB reduces ligand-independent N signaling activity. Our data suggest that lysosomal-associated functions regulated by the TFEB-V-ATPase axis might play a conserved role in shaping cell fate.

No MeSH data available.


Related in: MedlinePlus

Vha16-1 is required for in pupal SOPs differentiation. (A) Phenotypic defects associated with RNAi-mediated knockdown of Vha16-1 expression with the indicated RNAi line using the driver pannier-Gal4 (pnr) compared to control. Defects are rescued by concomitant expression of the RNAi lines and a RNAi-resistant Vha16-1-HA construct. The domain of pannier-Gal4 expression is delimited by arrowheads. (B) Quantification of the number of bristle/area (bristle density) relative to the experiment as in A. Statistical analysis is based on Kruskal Wallis Test with Dunn multiple comparison relative to control.
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f0006: Vha16-1 is required for in pupal SOPs differentiation. (A) Phenotypic defects associated with RNAi-mediated knockdown of Vha16-1 expression with the indicated RNAi line using the driver pannier-Gal4 (pnr) compared to control. Defects are rescued by concomitant expression of the RNAi lines and a RNAi-resistant Vha16-1-HA construct. The domain of pannier-Gal4 expression is delimited by arrowheads. (B) Quantification of the number of bristle/area (bristle density) relative to the experiment as in A. Statistical analysis is based on Kruskal Wallis Test with Dunn multiple comparison relative to control.

Mentions: Interestingly, expression of Vha16-1 RNAi in the pupal notum with pannier-Gal4 (pnr) led to a decrease in size of the adult thorax, which is formed by the fusion of the left and right nota. This phenotype is coupled with depigmentation and misorientation of bristles, a known effect in Drosophila of reduced V-ATPase and lysosomal activity6,37 (Fig. 6A). Importantly, the density of microchaeta, which derive from pupal SOPs, was also increased independent of thorax size (Fig. 6B), suggesting that reduction of Vha16-1 might weaken N -mediated lateral inhibition during pupal life. These effects are specific to depletion of Vha16-1, as they were rescued by concomitant overexpression of RNAi-resistant Vha16-1 tagged with HA (Vha16-1-HA, see Fig. S1F for details; Fig. 6A and B). Similar results were obtained by downregulating VhaPPA1-1, the gene encoding the component of the membrane-embedded V0 sector c” (Fig. S2I), as previously reported.38 However, Vha16-1 is not sufficient to promote ectopic PNC formation. In fact, compared to controls, overexpression of Vha16-1-HA per se in the wing pouch or notum did not perturb microcheta formation (Fig. 6B), suggesting that the patterning activity of Vha16-1 requires additional factors. Overall, these data indicate that Mitf and components of the V-ATPase might be functional elements of the proneural patterning machinery in wing discs epithelia.Figure 6.


Control of lysosomal biogenesis and Notch-dependent tissue patterning by components of the TFEB-V-ATPase axis in Drosophila melanogaster.

Tognon E, Kobia F, Busi I, Fumagalli A, De Masi F, Vaccari T - Autophagy (2016)

Vha16-1 is required for in pupal SOPs differentiation. (A) Phenotypic defects associated with RNAi-mediated knockdown of Vha16-1 expression with the indicated RNAi line using the driver pannier-Gal4 (pnr) compared to control. Defects are rescued by concomitant expression of the RNAi lines and a RNAi-resistant Vha16-1-HA construct. The domain of pannier-Gal4 expression is delimited by arrowheads. (B) Quantification of the number of bristle/area (bristle density) relative to the experiment as in A. Statistical analysis is based on Kruskal Wallis Test with Dunn multiple comparison relative to control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4836007&req=5

f0006: Vha16-1 is required for in pupal SOPs differentiation. (A) Phenotypic defects associated with RNAi-mediated knockdown of Vha16-1 expression with the indicated RNAi line using the driver pannier-Gal4 (pnr) compared to control. Defects are rescued by concomitant expression of the RNAi lines and a RNAi-resistant Vha16-1-HA construct. The domain of pannier-Gal4 expression is delimited by arrowheads. (B) Quantification of the number of bristle/area (bristle density) relative to the experiment as in A. Statistical analysis is based on Kruskal Wallis Test with Dunn multiple comparison relative to control.
Mentions: Interestingly, expression of Vha16-1 RNAi in the pupal notum with pannier-Gal4 (pnr) led to a decrease in size of the adult thorax, which is formed by the fusion of the left and right nota. This phenotype is coupled with depigmentation and misorientation of bristles, a known effect in Drosophila of reduced V-ATPase and lysosomal activity6,37 (Fig. 6A). Importantly, the density of microchaeta, which derive from pupal SOPs, was also increased independent of thorax size (Fig. 6B), suggesting that reduction of Vha16-1 might weaken N -mediated lateral inhibition during pupal life. These effects are specific to depletion of Vha16-1, as they were rescued by concomitant overexpression of RNAi-resistant Vha16-1 tagged with HA (Vha16-1-HA, see Fig. S1F for details; Fig. 6A and B). Similar results were obtained by downregulating VhaPPA1-1, the gene encoding the component of the membrane-embedded V0 sector c” (Fig. S2I), as previously reported.38 However, Vha16-1 is not sufficient to promote ectopic PNC formation. In fact, compared to controls, overexpression of Vha16-1-HA per se in the wing pouch or notum did not perturb microcheta formation (Fig. 6B), suggesting that the patterning activity of Vha16-1 requires additional factors. Overall, these data indicate that Mitf and components of the V-ATPase might be functional elements of the proneural patterning machinery in wing discs epithelia.Figure 6.

Bottom Line: In vertebrates, TFEB (transcription factor EB) and MITF (microphthalmia-associated transcription factor) family of basic Helix-Loop-Helix (bHLH) transcription factors regulates both lysosomal function and organ development.Similar to our findings in Drosophila, in human breast epithelial cells we observe that impairment of the Vha16-1 human ortholog ATP6V0C changes the size and function of the endolysosomal compartment and that depletion of TFEB reduces ligand-independent N signaling activity.Our data suggest that lysosomal-associated functions regulated by the TFEB-V-ATPase axis might play a conserved role in shaping cell fate.

View Article: PubMed Central - PubMed

Affiliation: a IFOM - FIRC Institute of Molecular Oncology , Milan , Italy.

ABSTRACT
In vertebrates, TFEB (transcription factor EB) and MITF (microphthalmia-associated transcription factor) family of basic Helix-Loop-Helix (bHLH) transcription factors regulates both lysosomal function and organ development. However, it is not clear whether these 2 processes are interconnected. Here, we show that Mitf, the single TFEB and MITF ortholog in Drosophila, controls expression of vacuolar-type H(+)-ATPase pump (V-ATPase) subunits. Remarkably, we also find that expression of Vha16-1 and Vha13, encoding 2 key components of V-ATPase, is patterned in the wing imaginal disc. In particular, Vha16-1 expression follows differentiation of proneural regions of the disc. These regions, which will form sensory organs in the adult, appear to possess a distinctive endolysosomal compartment and Notch (N) localization. Modulation of Mitf activity in the disc in vivo alters endolysosomal function and disrupts proneural patterning. Similar to our findings in Drosophila, in human breast epithelial cells we observe that impairment of the Vha16-1 human ortholog ATP6V0C changes the size and function of the endolysosomal compartment and that depletion of TFEB reduces ligand-independent N signaling activity. Our data suggest that lysosomal-associated functions regulated by the TFEB-V-ATPase axis might play a conserved role in shaping cell fate.

No MeSH data available.


Related in: MedlinePlus