Functional Specialization Among Members Of Knickkopf Family Of Proteins In Insect Cuticle Organization

Abstract

<div><p>Our recent study on the functional analysis of the Knickkopf protein from <i>T. castaneum</i> (TcKnk), indicated a novel role for this protein in protection of chitin from degradation by chitinases. Knk is also required for the laminar organization of chitin in the procuticle. During a bioinformatics search using this protein sequence as the query, we discovered the existence of a small family of three <i>Knk</i>-like genes (including the prototypical <i>TcKnk</i>) in the <i>T. castaneum</i> genome as well as in all insects with completed genome assemblies. The two additional <i>Knk</i>-like genes have been named <i>TcKnk2</i> and <i>TcKnk3</i>. Further complexity arises as a result of alternative splicing and alternative polyadenylation of transcripts of <i>TcKnk3</i>, leading to the production of three transcripts (and by inference, three proteins) from this gene. These transcripts are named <i>TcKnk3-Full Length</i> (<i>TcKnk3-FL</i>), <i>TcKnk3-5′</i> and <i>TcKnk3-3′</i>. All three <i>Knk</i>-family genes appear to have essential and non-redundant functions. RNAi for <i>TcKnk</i> led to developmental arrest at every molt, while down-regulation of either <i>TcKnk2</i> or one of the three <i>TcKnk3</i> transcripts (<i>TcKnk3</i>-3′) resulted in specific molting arrest only at the pharate adult stage. All three <i>Knk</i> genes appear to influence the total chitin content at the pharate adult stage, but to variable extents. While <i>TcKnk</i> contributes mostly to the stability and laminar organization of chitin in the elytral and body wall procuticles, proteins encoded by <i>TcKnk2</i> and <i>TcKnk3</i>-<i>3′</i> transcripts appear to be required for the integrity of the body wall denticles and tracheal taenidia, but not the elytral and body wall procuticles. Thus, the three members of the Knk-family of proteins perform different essential functions in cuticle formation at different developmental stages and in different parts of the insect anatomy.</p></div