WHY ARE 2 DIFFERENT CROSS-LINKERS NECESSARY FOR ACTIN BUNDLE FORMATION IN-VIVO AND WHAT DOES EACH CROSS-LINK CONTRIBUTE

In developing Drosophila bristles two species of cross-linker, the for ked proteins and fascin, connect adjacent actin filaments into bundles . Bundles form in three phases: (a) tiny bundles appear; (b) these bun dles aggregate into larger bundles; and (c) the filaments become maxim ally cross-linked by fascin. In mutants that completely lack forked, a ggregation of the bundles does not occur so that the mature bundles co nsist of <50 filaments versus similar to 700 for wild type. If the for ked concentration is genetically reduced to half the wild type, aggreg ation of the tiny bundles occurs but the filaments are poorly ordered albeit with small patches of fascin cross-linked filaments. In mutants containing an excess of forked, all the bundles tend to aggregate and the filaments are maximally crossbridged by fascin. Alternatively, if fascin is absent, phases 1 and 2 occur normally but the resultant bun dles are twisted and the filaments within them are poorly ordered. By extracting fully elongated bristles with potassium iodide which remove s fascin but leaves forked, the bundles change from being straight to twisted and the filaments within them become poorly ordered. From thes e observations we conclude that (a) forked is used early in developmen t to aggregate the tiny bundles into larger bundles; and (b) forked fa cilitates fascin entry into the bundles to maximally cross-link the ac tin filaments into straight, compact, rigid bundles. Thus, forked alig ns the filaments and then directs fascin binding so that inappropriate cross-linking does not occur.