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The Homeodomain Iroquois Proteins Control Cell Cycle Progression and Regulate the Size of Developmental Fields.

Barrios N, González-Pérez E, Hernández R, Campuzano S - PLoS Genet. (2015)

Bottom Line: Conversely, their increased expression causes cell-cycle arrest, down-regulating the activity of the Cyclin E/Cdk2 complex.Thus, Drosophila Iroquois proteins are able to regulate cell-autonomously the growth of the territories they specify.Moreover, our results provide a molecular mechanism for a role of Iroquois/Irx genes as tumour suppressors.

View Article: PubMed Central - PubMed

Affiliation: Department of Development and Differentiation, Centro de Biología Molecular Severo Ochoa, CSIC-UAM, Madrid, Spain.

ABSTRACT
During development, proper differentiation and final organ size rely on the control of territorial specification and cell proliferation. Although many regulators of these processes have been identified, how both are coordinated remains largely unknown. The homeodomain Iroquois/Irx proteins play a key, evolutionarily conserved, role in territorial specification. Here we show that in the imaginal discs, reduced function of Iroquois genes promotes cell proliferation by accelerating the G1 to S transition. Conversely, their increased expression causes cell-cycle arrest, down-regulating the activity of the Cyclin E/Cdk2 complex. We demonstrate that physical interaction of the Iroquois protein Caupolican with Cyclin E-containing protein complexes, through its IRO box and Cyclin-binding domains, underlies its activity in cell-cycle control. Thus, Drosophila Iroquois proteins are able to regulate cell-autonomously the growth of the territories they specify. Moreover, our results provide a molecular mechanism for a role of Iroquois/Irx genes as tumour suppressors.

No MeSH data available.


Related in: MedlinePlus

Cell-autonomous increase in cell proliferation in iro mutants.Lateral (A, C) and dorsal (B, D) views of heads of flies of the indicated genotypes. (E- F’) Expression of Wg (green) and Phalloidin staining (red) in wild-type (E, E’) and eyGal4>mirr RNAi (two copies of mirr RNAi, flies raised at 29°C, F, F’) eye discs. (E and E’ and F and F’ are different focal planes of the same disc). Arrowheads and arrow mark the position of the morphogenetic furrow. (G-K) Mitotic patterns (phospho-Histone H3 staining, green, G, H; red I, J) and quantification of the relative mitotic index (K) in Iro-expressing territories (white dotted areas in G-J) and in the prospective proximal notum (yellow dotted areas in I, J). (*p<0.05; **p<0.005). (L) G1/S transition is accelerated in iro mutant cells. Representative profiles of FACS analysis of cells dissociated from iroDFM3/iroGal4 UAS-GFP wing discs. (The differences in the percentages of G1 and (G2+S) cells between the GFP+ and GFP- populations are statistically significant, **p<0.005). (M, N) Reduction of Mirr levels (one copy of UAS-mirr RNAi, larvae raised at 25°C) and over-expression of stg synergistically interact to increase eye size. (M) Quantification of the fraction of enlarged eyes in flies of the indicated genotypes (average from two independent experiments, n>100, *p<0.05). (N) Representative mutant enlarged eye. In this and following figures, the eye discs are oriented dorsal up and posterior to the right, and the wing discs, ventral up and posterior to the right. Quantitative data are shown as arithmetic mean +/- SD (error bars). WT, wild-type.
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pgen.1005463.g001: Cell-autonomous increase in cell proliferation in iro mutants.Lateral (A, C) and dorsal (B, D) views of heads of flies of the indicated genotypes. (E- F’) Expression of Wg (green) and Phalloidin staining (red) in wild-type (E, E’) and eyGal4>mirr RNAi (two copies of mirr RNAi, flies raised at 29°C, F, F’) eye discs. (E and E’ and F and F’ are different focal planes of the same disc). Arrowheads and arrow mark the position of the morphogenetic furrow. (G-K) Mitotic patterns (phospho-Histone H3 staining, green, G, H; red I, J) and quantification of the relative mitotic index (K) in Iro-expressing territories (white dotted areas in G-J) and in the prospective proximal notum (yellow dotted areas in I, J). (*p<0.05; **p<0.005). (L) G1/S transition is accelerated in iro mutant cells. Representative profiles of FACS analysis of cells dissociated from iroDFM3/iroGal4 UAS-GFP wing discs. (The differences in the percentages of G1 and (G2+S) cells between the GFP+ and GFP- populations are statistically significant, **p<0.005). (M, N) Reduction of Mirr levels (one copy of UAS-mirr RNAi, larvae raised at 25°C) and over-expression of stg synergistically interact to increase eye size. (M) Quantification of the fraction of enlarged eyes in flies of the indicated genotypes (average from two independent experiments, n>100, *p<0.05). (N) Representative mutant enlarged eye. In this and following figures, the eye discs are oriented dorsal up and posterior to the right, and the wing discs, ventral up and posterior to the right. Quantitative data are shown as arithmetic mean +/- SD (error bars). WT, wild-type.

Mentions: We found that iroEGP1 homozygous flies and those harbouring the iroEGP1 allele combined with a deficiency of the whole Iro-C (iroDFM3, S1A Fig) had dorsally enlarged eyes (Fig 1A–1D, 5% of iroEGP1 flies, 36% of the iroEGP1 /iroDFM3 everted flies). The cephalic capsule was morphologically normal, except for alterations in the number of orbital bristles (Fig 1D, arrowhead). In third instar wild-type eye imaginal discs, the three Iro genes are expressed in a dorsal domain ahead of the morphogenetic furrow (S1B and S1C Fig, see also [10, 14]). In contrast, in iroEGP1 /iroDFM3 eye discs the expression of caup was undetectable and that of ara was strongly decreased, while mirr expression was not affected (S1D–S1F Fig). Dorsally enlarged eyes were also found in 51% of the flies depleted of Mirr (by expression of two copies of UAS-mirr RNAi driven by eyGal4 at 25°C).


The Homeodomain Iroquois Proteins Control Cell Cycle Progression and Regulate the Size of Developmental Fields.

Barrios N, González-Pérez E, Hernández R, Campuzano S - PLoS Genet. (2015)

Cell-autonomous increase in cell proliferation in iro mutants.Lateral (A, C) and dorsal (B, D) views of heads of flies of the indicated genotypes. (E- F’) Expression of Wg (green) and Phalloidin staining (red) in wild-type (E, E’) and eyGal4>mirr RNAi (two copies of mirr RNAi, flies raised at 29°C, F, F’) eye discs. (E and E’ and F and F’ are different focal planes of the same disc). Arrowheads and arrow mark the position of the morphogenetic furrow. (G-K) Mitotic patterns (phospho-Histone H3 staining, green, G, H; red I, J) and quantification of the relative mitotic index (K) in Iro-expressing territories (white dotted areas in G-J) and in the prospective proximal notum (yellow dotted areas in I, J). (*p<0.05; **p<0.005). (L) G1/S transition is accelerated in iro mutant cells. Representative profiles of FACS analysis of cells dissociated from iroDFM3/iroGal4 UAS-GFP wing discs. (The differences in the percentages of G1 and (G2+S) cells between the GFP+ and GFP- populations are statistically significant, **p<0.005). (M, N) Reduction of Mirr levels (one copy of UAS-mirr RNAi, larvae raised at 25°C) and over-expression of stg synergistically interact to increase eye size. (M) Quantification of the fraction of enlarged eyes in flies of the indicated genotypes (average from two independent experiments, n>100, *p<0.05). (N) Representative mutant enlarged eye. In this and following figures, the eye discs are oriented dorsal up and posterior to the right, and the wing discs, ventral up and posterior to the right. Quantitative data are shown as arithmetic mean +/- SD (error bars). WT, wild-type.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4549242&req=5

pgen.1005463.g001: Cell-autonomous increase in cell proliferation in iro mutants.Lateral (A, C) and dorsal (B, D) views of heads of flies of the indicated genotypes. (E- F’) Expression of Wg (green) and Phalloidin staining (red) in wild-type (E, E’) and eyGal4>mirr RNAi (two copies of mirr RNAi, flies raised at 29°C, F, F’) eye discs. (E and E’ and F and F’ are different focal planes of the same disc). Arrowheads and arrow mark the position of the morphogenetic furrow. (G-K) Mitotic patterns (phospho-Histone H3 staining, green, G, H; red I, J) and quantification of the relative mitotic index (K) in Iro-expressing territories (white dotted areas in G-J) and in the prospective proximal notum (yellow dotted areas in I, J). (*p<0.05; **p<0.005). (L) G1/S transition is accelerated in iro mutant cells. Representative profiles of FACS analysis of cells dissociated from iroDFM3/iroGal4 UAS-GFP wing discs. (The differences in the percentages of G1 and (G2+S) cells between the GFP+ and GFP- populations are statistically significant, **p<0.005). (M, N) Reduction of Mirr levels (one copy of UAS-mirr RNAi, larvae raised at 25°C) and over-expression of stg synergistically interact to increase eye size. (M) Quantification of the fraction of enlarged eyes in flies of the indicated genotypes (average from two independent experiments, n>100, *p<0.05). (N) Representative mutant enlarged eye. In this and following figures, the eye discs are oriented dorsal up and posterior to the right, and the wing discs, ventral up and posterior to the right. Quantitative data are shown as arithmetic mean +/- SD (error bars). WT, wild-type.
Mentions: We found that iroEGP1 homozygous flies and those harbouring the iroEGP1 allele combined with a deficiency of the whole Iro-C (iroDFM3, S1A Fig) had dorsally enlarged eyes (Fig 1A–1D, 5% of iroEGP1 flies, 36% of the iroEGP1 /iroDFM3 everted flies). The cephalic capsule was morphologically normal, except for alterations in the number of orbital bristles (Fig 1D, arrowhead). In third instar wild-type eye imaginal discs, the three Iro genes are expressed in a dorsal domain ahead of the morphogenetic furrow (S1B and S1C Fig, see also [10, 14]). In contrast, in iroEGP1 /iroDFM3 eye discs the expression of caup was undetectable and that of ara was strongly decreased, while mirr expression was not affected (S1D–S1F Fig). Dorsally enlarged eyes were also found in 51% of the flies depleted of Mirr (by expression of two copies of UAS-mirr RNAi driven by eyGal4 at 25°C).

Bottom Line: Conversely, their increased expression causes cell-cycle arrest, down-regulating the activity of the Cyclin E/Cdk2 complex.Thus, Drosophila Iroquois proteins are able to regulate cell-autonomously the growth of the territories they specify.Moreover, our results provide a molecular mechanism for a role of Iroquois/Irx genes as tumour suppressors.

View Article: PubMed Central - PubMed

Affiliation: Department of Development and Differentiation, Centro de Biología Molecular Severo Ochoa, CSIC-UAM, Madrid, Spain.

ABSTRACT
During development, proper differentiation and final organ size rely on the control of territorial specification and cell proliferation. Although many regulators of these processes have been identified, how both are coordinated remains largely unknown. The homeodomain Iroquois/Irx proteins play a key, evolutionarily conserved, role in territorial specification. Here we show that in the imaginal discs, reduced function of Iroquois genes promotes cell proliferation by accelerating the G1 to S transition. Conversely, their increased expression causes cell-cycle arrest, down-regulating the activity of the Cyclin E/Cdk2 complex. We demonstrate that physical interaction of the Iroquois protein Caupolican with Cyclin E-containing protein complexes, through its IRO box and Cyclin-binding domains, underlies its activity in cell-cycle control. Thus, Drosophila Iroquois proteins are able to regulate cell-autonomously the growth of the territories they specify. Moreover, our results provide a molecular mechanism for a role of Iroquois/Irx genes as tumour suppressors.

No MeSH data available.


Related in: MedlinePlus