All filtering was performed at each solitary Z-slice. NIHMS564262-supplement-Supp_Movie_S3.AVI (1.4M) GUID:?37F9C6CE-F4B3-4980-B051-7B4FC2E745F8 Supp Movie S4: Movie 4. Inner bud cells showing lower motility. This fluorescence time-lapse movie acquired under the same conditions as the additional movies shows slower average motility of cells photo-converted in a region of the inner bud. Remaining: reddish; center: green; right: overlay. NIHMS564262-supplement-Supp_Movie_S4.AVI (14M) GUID:?0E845A28-667A-45CD-B4B4-E6BE2783FAB7 Supp Movie S5: Movie 5. Outer bud cells inside a salivary gland treated with antibodies against 6 and 1 integrin antibodies at 100 g/ml each. Notice the loss of association of the migrating cells with the basement membrane. Remaining: reddish; center: green; right: overlay. NIHMS564262-supplement-Supp_Movie_S5.AVI (7.0M) GUID:?F1E27C3B-0A00-4101-B0B2-6818D229C33A Supp Movie S6: Movie 6. Composite movie showing effects of inhibiting integrins on outer bud cell motility rates and patterns. The left panel shows untreated control inner bud cells, and the right panel shows inner bud cells treated with 100 g/ml inhibitory anti-6 and -1 integrin antibodies (Integrin I.). Both panels display overlay of reddish and green channels. Notice the loss of cell association with the basement membrane after integrin inhibition, as well as the decreased average velocity of migration of the cells, reflected in the reduced lateral dispersion of cells along the basement membrane. NIHMS564262-supplement-Supp_Movie_S6.AVI (11M) GUID:?84C80447-2F31-4143-B9B3-8C3C2F0227FE Supp Movie S7: Movie 7. Outer bud cells inside a salivary gland treated with inhibitory anti-E-cadherin monoclonal antibody (100 g/ml ECCD-1). Cinnamyl alcohol Notice the continued association of some of the cells with the basement membrane. Remaining: reddish; center: green; right: overlay. NIHMS564262-supplement-Supp_Movie_S7.AVI (7.4M) GUID:?4BA2C298-0B8F-4F3E-92D5-72EF086BDA0F Supp Movie S8: Movie 8. Composite movie showing effects of inhibiting E-cadherin on inner and outer bud cells of salivary glands. The top panels show untreated control inner and outer bud cells, and the lower panels show these cells in glands treated with inhibitory anti-E-cadherin monoclonal antibody (100 g/ml ECCD-1, labeled Ecad). For clearer visualization of the reddish photo-converted cells, the remaining panels show only the reddish channel in grayscale, while the ideal panels show overlay of the reddish and green channels. Notice the disruption of cell-cell adhesion and the formation of black gaps or holes in the inner bud epithelium. NIHMS564262-supplement-Supp_Movie_S8.AVI (21M) GUID:?E082C289-309E-4D1B-81D5-4BCE29047BFC Supp Movie S9: Movie 9. Outer bud cells inside a salivary gland treated with 50 M blebbistatin. Notice the continued association of some of the cells with the basement membrane. Remaining: basement membrane stained for collagen IV; center: photo-converted outer bud cells; right: overlay of collagen IV (pseudo-colored green) and reddish channel showing photo-converted KikGR outer bud cells. The collagen IV was imaged using a 642 nm Cinnamyl alcohol laser, because the green channel cannot be used in order to avoid blebbistatin photo-inactivation and resultant cytotoxicity. NIHMS564262-supplement-Supp_Movie_S9.AVI (3.4M) GUID:?131B3BDA-99EF-4E47-8BAE-A78E4A764D04 Supp Movie S10: Movie 10. Composite movie comparing control salivary gland with glands treated with monoclonal antibody inhibitors of E-cadherin (Ecad) or integrin 6 and 1 subunits (Integrin), or with blebbistatin to inhibit myosin II isoforms (Myosin II). For all four panels, reddish shows photo-converted cells, and green shows non-photo-converted KikGR-expressing cells in panels ACC. In panel D, the basement membrane visualized using a collagen IV antibody imaged at 642 nm and pseudo-colored green to avoid blebbistatin degradation and cytotoxicity. NIHMS564262-supplement-Supp_Movie_S10.AVI (7.6M) GUID:?5CE697DA-55E0-4147-9E93-FC57D154F9B9 Abstract Background Epithelial cells of developing embryonic organs, such as salivary glands, can display considerable motility during branching morphogenesis. Their dynamic motions and molecules involved in their migration are not fully characterized. Results We Rabbit Polyclonal to KAP1 generated transgenic mice expressing photo-convertible KikGR and tracked the motions of individual cells highlighted by reddish fluorescence in different regions of developing salivary glands. Motility was highest for outer bud epithelial cells adjacent to the basement membrane, reduced inner bud cells, and least expensive in duct cells. The highly motile outer cells contacting the basement membrane were pleomorphic, whereas inner cells were rounded. Peripheral cell motility was disrupted Cinnamyl alcohol by antibodies inhibiting 6+1 integrins and the non-muscle myosin II inhibitor blebbistatin. Inner bud cell migration was unaffected by these inhibitors, but their rate of migration was stimulated by inhibiting E-cadherin. Conclusions Cell motility in developing salivary glands was highest in cells in contact with the basement membrane. The basement membrane-associated motility of these outer bud cells depended.