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Fig. 4. Drosophila Cas and integrin mutants exhibit SNa motor axon defasciculation defects. (A-J) Immunostaining as in Fig. 3. The intersegmental nerve (ISN) and the dorsal (D) and lateral (L) branch of segmental nerve a (SNa) are indicated. (A) In DCas^{Df(3L)Exel6083}/+ embryos, axons within the SNa defasciculate normally from the SN and extend past the ventral musculature as a single tightly fasciculated bundle. Near the dorsal edge of muscle 12, SNa axons defasciculate (small arrow) to generate a dorsal (D) and a lateral (L) branch. Axons within the dorsal branch extend dorsally between muscles 22 and 23 and then make two characteristic turns (open arrows), continuing dorsally between muscles 23 and 24. (B-F) DCas loss-of-function mutant embryos often fail to reach their muscle targets and exhibit a range of SNa axon guidance phenotypes indicative of defects in axon-axon defasciculation. (B,C) SNa axons in DCas mutants exhibit abnormal fasciculation (arrowheads) of the lateral branch with the dorsal branch (B, arrow), and also of the dorsal branch with the lateral branch (C, arrow). (D,E) In DCas mutants, axons within the dorsal branch of the SNa often do not make their two characteristic turns, extending past their defasciculation choice point (arrow, D) or stalling in the vicinity of this choice point (arrow, E). (F) SNa axons in DCas mutants are also abnormally fasciculated, extending past their defasciculation point (arrow) but eventually taking an abnormal route to reach lateral muscle targets (open arrows). (G,H) SNa axon guidance defects seen in 1 integrin (mew^M6) (G, 1Int) and 2 integrin (if^K27E) (H, 2Int) LOF mutants resemble those observed in DCas LOF mutants, including: SNa stalling past the dorsal SNa choice point (large arrow, G), axons fused with the ISN (arrowhead, H), and SNa axons within the dorsal branch growing past their choice point (large arrow, H). (I,J) Expression of DCas rescues SNa defects observed in DCas LOF mutants. Expression of either ^MycDCas under the DCas^P1-GAL4 driver (I, UAS-^MycDCas/+; Df(3L)ED201/DCas^P1) or ^MycDCas in all neurons using the ELAV-GAL4 driver (J, ELAV-GAL4/UAS-^MycDCas; DCas^{Df(3L)Exel6083}/DCas^{Df(3L)Exel6083}) restores the normal SNa trajectory and innervation pattern in DCas mutants (open arrows). Scale bar: 15 µm. (K) Schematic summary of SNa motor axon guidance phenotypes.

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