Fig 3 - uploaded by John Pearson
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timp over-expression impairs cyst encapsulation without affecting cell fate acquisition. (A, B) Ovarioles from control (no heat-shock) and experimental (daily heat-shock regime for 2 weeks post-eclosion) animals carrying UAS-timp and hs-Gal4. Arrows compare a normal egg chamber containing 16 germ cells with an abnormal, fused egg chamber encasing 32 germ cells. (C, D) Control (c587-Gal4) and experimental (c587-Gal4; UAS-timp) ovarioles grown at 18°C and then switched to 25°C for 9 days. Arrowheads indicate Lamin C-positive stalk cells. (E, F) Control (c587-Gal4) and experimental (c587-Gal4; UAS-timp) ovarioles grown at 18°C and aged 4 weeks at 25°C. Large, empty arrowheads show Mmp-1 accumulation from region 2. The ring of strong Mmp1 staining that coincides with the region contracting to pinch off a new egg chamber in controls is absent in timp-overexpressing germaria (small, empty arrowheads). Occasionally, the TF of experimental germaria show a prominent accumulation of Mmp1 staining, indicating that timp overexpression may affect Mmp1 localization in this region. Images can be composites of several focal planes.
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Context 1
... absence of gross alterations in the localization of ECM proteins in mutant germaria could suggest that changes to MMP activity resulting from the lack of timp are highly localized (e.g. affecting only specific cell types) and/or modify ECM properties without affecting its overall distribution. Overexpression of timp in the somatic component of otherwise wild-type ovaries using a UASt-timp insertion [28] combined with either a heat-shock Gal4 or the germarium- specific Gal4 c587 [29] resulted in the formation of compound follicles containing more than one germline cyst (Fig 3A and 3B). This fusion phenotype could be caused by a defect in the encapsulation of the cysts in germarial Region 2 or by a failure in the specification of stalk cells that separate adjacent egg chambers [30]. Significantly, c587-Gal4 expression, which pro- duced the strongest UASt-timp induced phenotypes, is specific to a subset of somatic cells in the germarium [29], implying the need for a fine regulation of extracellular proteolytic activity in anterior ovarian tissues during early cyst formation. Examination of ovaries containing timp-induced egg chamber fusions revealed the presence of cells expressing the stalk cell marker Lamin C towards the periphery of the chambers, in the approximate position where stalk cells would normally be located (Fig 3C and 3D). These observations emphasize the importance of the fine-regulation of timp function in the somatic cells of the germarium for the correct organization of stalk cells and for the encapsulation of new cysts, a process that requires extensive cell migration and ECM remodeling. Importantly, enlarged germaria induced by timp overexpression lack the strong accumulation of Mmp1 in the region where the first interfollicular stalk would form (Fig 3E and 3F; see also Fig 7 for a detailed description of Mmp1 localization), supporting the idea that timp overexpression may affect Mmp1 activ- ity during cyst ...
Context 2
... absence of gross alterations in the localization of ECM proteins in mutant germaria could suggest that changes to MMP activity resulting from the lack of timp are highly localized (e.g. affecting only specific cell types) and/or modify ECM properties without affecting its overall distribution. Overexpression of timp in the somatic component of otherwise wild-type ovaries using a UASt-timp insertion [28] combined with either a heat-shock Gal4 or the germarium- specific Gal4 c587 [29] resulted in the formation of compound follicles containing more than one germline cyst (Fig 3A and 3B). This fusion phenotype could be caused by a defect in the encapsulation of the cysts in germarial Region 2 or by a failure in the specification of stalk cells that separate adjacent egg chambers [30]. Significantly, c587-Gal4 expression, which pro- duced the strongest UASt-timp induced phenotypes, is specific to a subset of somatic cells in the germarium [29], implying the need for a fine regulation of extracellular proteolytic activity in anterior ovarian tissues during early cyst formation. Examination of ovaries containing timp-induced egg chamber fusions revealed the presence of cells expressing the stalk cell marker Lamin C towards the periphery of the chambers, in the approximate position where stalk cells would normally be located (Fig 3C and 3D). These observations emphasize the importance of the fine-regulation of timp function in the somatic cells of the germarium for the correct organization of stalk cells and for the encapsulation of new cysts, a process that requires extensive cell migration and ECM remodeling. Importantly, enlarged germaria induced by timp overexpression lack the strong accumulation of Mmp1 in the region where the first interfollicular stalk would form (Fig 3E and 3F; see also Fig 7 for a detailed description of Mmp1 localization), supporting the idea that timp overexpression may affect Mmp1 activ- ity during cyst ...
Context 3
... absence of gross alterations in the localization of ECM proteins in mutant germaria could suggest that changes to MMP activity resulting from the lack of timp are highly localized (e.g. affecting only specific cell types) and/or modify ECM properties without affecting its overall distribution. Overexpression of timp in the somatic component of otherwise wild-type ovaries using a UASt-timp insertion [28] combined with either a heat-shock Gal4 or the germarium- specific Gal4 c587 [29] resulted in the formation of compound follicles containing more than one germline cyst (Fig 3A and 3B). This fusion phenotype could be caused by a defect in the encapsulation of the cysts in germarial Region 2 or by a failure in the specification of stalk cells that separate adjacent egg chambers [30]. Significantly, c587-Gal4 expression, which pro- duced the strongest UASt-timp induced phenotypes, is specific to a subset of somatic cells in the germarium [29], implying the need for a fine regulation of extracellular proteolytic activity in anterior ovarian tissues during early cyst formation. Examination of ovaries containing timp-induced egg chamber fusions revealed the presence of cells expressing the stalk cell marker Lamin C towards the periphery of the chambers, in the approximate position where stalk cells would normally be located (Fig 3C and 3D). These observations emphasize the importance of the fine-regulation of timp function in the somatic cells of the germarium for the correct organization of stalk cells and for the encapsulation of new cysts, a process that requires extensive cell migration and ECM remodeling. Importantly, enlarged germaria induced by timp overexpression lack the strong accumulation of Mmp1 in the region where the first interfollicular stalk would form (Fig 3E and 3F; see also Fig 7 for a detailed description of Mmp1 localization), supporting the idea that timp overexpression may affect Mmp1 activ- ity during cyst ...
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