FIG. 2.

(A) Northern blot analysis of JAM4. A uniformly labeled probe corresponding to the full length of JAM4 was prepared. Blots with 20 μg of total RNA from each mouse tissue were hybridized with the probe (3,600,000 cpm) and exposed for 8 days. The mobilities of molecular mass standards (in kilobases) are indicated at the left. (B) Western blot with the anti-JAM4 antibody of rat kidney, intestine, spleen, and MTD-1A cells. Homogenates (total protein, 20 μg) from rat kidney, small intestine, spleen, and MTD-1A cells were resolved by SDS-PAGE and immunoblotted with the affinity-purified antibody against JAM4. Lane 1, spleen cells; lanes 2 and 4, kidney cells; lanes 3 and 5, small intestine cells; lanes 6 and 7, MTD-1A cells. For lanes 4, 5, and 7, the antibody was preincubated with 5 μM immunogen. The mobilities of molecular mass standards (in kilodaltons) are indicated at left. (C) FLAG-JAM4 in various cells. (a) Homogenates (total protein, 20 μg) from various stable transformants expressing FLAG-JAM4 were resolved by SDS-PAGE and immunoblotted with the anti-FLAG antibody. (b) Homogenates of stable transformants were treated with N-glycosidase F and charged onto SDS-PAGE gel with the in vitro transcription and translation product. The lane for the in vitro transcription and translation product was separately analyzed with the image analyzer. The remaining lanes were immunoblotted with the anti-FLAG antibody. The mobilities of molecular mass standards (in kilodaltons) are indicated at the left.