Fig. 3

jam3b expression in pigment cells and other cell types of post-embryonic zebrafish. (A) RT-PCR for jam3b and related 2-Ig domain receptor gene igsf11 in isolated melanophores (mel), xanthophores (xan), iridophores (irid), and whole fin tissue. –, no template control. pmela marks melanophores, colony stimulating factor 1 receptor a (csf1ra) marks xanthophores, and purine nucleoside phosphorylase 4a (pnp4a) most strongly marks iridophores and melanophores (Lang et al., 2009; Patterson and Parichy, 2013; Saunders et al., 2019). (B) jam3b and jam2a expression in melanophores, xanthophores, iridophores, and post-embryonic pigment progenitor cells (N ​= ​3954 ​cells total) by single cell RNA-sequencing across states of maturation during adult pigment pattern formation. Cells in which transcripts were not detected (nd) are shown in light grey. Numbers show percentages of cells within each cluster in which each transcript was detected Sparsity of transcript detection may represent biological heterogenities or stochastic dropout associated with imperfect capture or other aspects of scRNA-Seq methodology, as is typical for genes expressed at moderate or low levels (Qiu, 2020). igsf11 transcripts were expressed at levels too low to detect by scRNA-Seq (Eom et al., 2012). (C–E) Expression of jam3b:Jam3b-mCherry. (C) Upper, Segmental expression (arrowheads) associated with vasculature (arrowheads) in a late larva (10 SSL). Diffuse expression is also evident in fin (f). ∗, highly reflective melanoleucophores (Lewis et al., 2019) of the dorsal fin, not representative of Jam3b-mCherry expression. Lower left, Expression in an earlier larva (6 SSL), with labeling of vasculature (v) as well as myocytes (myo). Arrow, early larval melanophore at horizontal myoseptum. Lower right, Jam3b-mCherry co-expression with endothelial marker kdrl:GFP (lower right). Additional Jam3b-mCherry was evident in presumptive Schwann cells of an adjacent nerve fiber (n) (Ebnet, 2017; Scheiermann et al., 2007). (D) Within mature stripes, Jam3b-mCherry accumulated at sites of apparent overlap between melanophores but typically did not extend to where melanophore membranes bordered the interstripe (arrowheads). Different regions are shown of the same fish (14 SSL) after partial contraction of melanin granules with epinephrine, using widefield epifluorescence to capture tissue context. (E) Low and high magnification images of interstripe iridophores (7 and 6 SSL, respectively), in which Jam3b-mCherry accumulated at sites of juxtaposition between cells (arrows). Yellow dots outline individual iridophores for clarity at right. Scale bars, 500 ​μM, 50 ​μM, 20 ​μm (C), 50 ​μM (D), 20 ​μm (E).