Fig. 4. Mapping of the lamin Dm₀ interaction domain with JIL-1. (A) The truncated COOH-terminal lamin Dm₀ GST-fusion protein constructs used for domain mapping. The Ig-like fold of the lamin Dm₀ tail domain (white) is shown in black. Some of the constructs also contained parts of the rod domain (shown in gray). (B) S2 cell lysate was incubated with the various truncated lamin GST-fusion protein constructs shown in A or with a beads-only control and pelleted with glutathione-agarose beads. Interacting protein(s) were fractionated by SDS-PAGE, western blotted and probed with JIL-1 antiserum. Unincubated S2 cell lysate was included as a control (lane 6). The lamin-D1 and lamin-D3 constructs were able to pull down the 160 kDa JIL-1 protein (lanes 1 and 3) also detected in the cell lysate (lane 6), whereas no interaction was observed with the lamin-D2 and lamin-D4 constructs (lanes 2 and 4) or with the beads-only control (lane 5). This defined the lamin-D3 domain in the tail region as sufficient for mediating interactions with JIL-1. (C) Immunoblot of the input GST-fusion proteins used for the pull-down experiments in B detected with the anti-GST mAb 8C7. The relative migration of molecular weight markers is indicated to the right in kDa.