Live imaging of Drosophila gonad formation reveals roles for Six4 in regulating germline and somatic cell migration

Additional file 1:

PGC migration to the gonads. An embryo expressing eGFP-Vasa in primordial germ cells imaged during stages 11–14. Germ cells move to the mesoderm by a variety of routes then coalesce with SGPs (not visible) following germ band retraction. The gonad then compacts to a spherical shape. Image stacks were collected at 90s intervals over 12 hours of embryonic development. The video was made from maximum projections of data from 30 deconvolved z sections at each time point taken at 1 μm intervals. The frame rate is 29.97 fps. Still images derived from these data are shown in Figure 1.

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Additional file 2:

Somatic cell movements during gonadogenesis. Male wild type embryo carrying a Six4-nls-eGFP transgene imaged during stages 12–14. The three SGP clusters originating in parasegments 10–12 associate during germ band retraction before further SGP movements result in compaction of the gonad. The msSGPs migrate anteriorly and dorsally to join the gonad. Image stacks were captured at 90s intervals. The video was made from maximum projections of data from 26 deconvolved z sections at each time point, taken at 1 μm intervals. The frame rate is 29.97 fps. Still images derived from these data are shown in Figure 2a

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Additional file 3:

SGP movements in an Hmgcr homozygous mutant embryo. Female Hmgcr homozygous embryo carrying a Six4-nls-eGFP transgene imaged during stages 12–14. The Hmgcr mutation allows SGP movements to be observed in the absence of associating PGCs. The three SGP clusters (red arrows) are in close proximity by mid stage 12 and remain associated for some time before the initiation of gonad compaction. The video was made from maximum projections of data from 26 deconvolved z sections taken at 1 μm intervals. Image stacks were captured at 90s intervals. The frame rate is 29.97 fps. Still images derived from these data are shown in Figure 2b.

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Additional file 4:

Cells migrate towards the gonad from a ventral, posterior position in female embryos. Wild type female embryo carrying a Six4-nls-eGFP transgene. A small number of cells migrate to the gonad from parasegment 13 in a similar manner to the msSGPs observed in males. Some remain associated with the gonad while others move rapidly away. Images were captured at 2 minute intervals. The video was made from a single z section at each time point. The frame rate is 29.97 fps. Still images derived from this data are shown in Figure 2c.

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Additional file 5:

Six4 is required for PGC migration to the SGPs. Six4¹³¹ homozygous mutant embryo expressing eGFP-Vasa in PGCs. A small number of PGCs (white arrow) become associated with SGPs in the normal location of the gonad following germ band retraction. The majority remain in ventral and posterior positions at this time. The video was made from maximum projections of data from 30 deconvolved z sections 1 μm apart. Image stacks were collected at 90s intervals. The frame rate is 29.97 fps. Still images derived from these data are shown in Figure 3c.

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Additional file 6:

Six4 is required for SGPs to form a unified gonad. Six4¹³¹ homozygous mutant embryo carrying a Six4-nls-eGFP transgene. The SGP clusters derived from parasegments 11 and 12 (cyan arrows) associate and compact to form a gonad-like structure in an abnormally posterior position. The cluster derived from parasegment 10 (red arrows) remains isolated but these cells also compact to form a tight cluster. The video was made from maximum projections of data from 26 deconvolved z sections 1 μm apart. The frame rate is 29.97 fps. Image stacks were collected at 90s intervals. Still images derived from these data are shown in Figure 4.

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