A Novel Collapsing Response Mediator Protein 2 Binding Domain for Kinesin Light Chain 3

Significant similarity exists between primary cilia and sperm flagella. Both contain an axoneme and both require presence of ODF2. Therefore, primary cilia can be used as a partial model to study the assembly of sperm flagella. Recently, we have shown that Collapsin Response Mediator Protein 2 is required for ciliogenesis, localizes to the basal body, and can be transported into the primary cilium, however, the involved mechanism is not clear, but was proposed to involve kinesin proteins. KLC3 is the only known kinesin light chain expressed in elongating spermatids that develop the flagellum. In this study, we investigated the expression of CRMP2 in mouse testis and we studied if CRMP2 and KLC3 interact. We observed CRMP2 mRNA expression in testis, along with expression of the other family members CRMP1, CRMP3 and CRMP4. CRMP2 protein localized in elongating spermatids to the neck region containing the centrioles, structures that also form the basal body. We show interaction of KLC3 and CRMP2 in cell culture and discovered that the interaction requires a sequence of CRMP2 different from that which is known to bind KLC1. We also show that KLC3 expression can change the localization of CRMP2 from one associated with microtubules to one associated with complexes of KLC3 and mitochondria. Our study suggests a role for CRMP2 in the elongating spermatid

Tubulin lattice in cilia is in a stressed form regulated by microtubule inner proteins

Cilia, the hair-like protrusions that beat at high frequencies to propel a cell or move fluid around are composed of radially bundled doublet microtubules. In this study, we present a near-atomic resolution map of the Tetrahymena doublet microtubule by cryoelectron microscopy. The map demonstrates that the network of microtubule inner proteins weaves into the tubulin lattice and forms an inner sheath. From mass spectrometry data and de novo modeling, we identified Rib43a proteins as the filamentous microtubule inner proteins in the protofilament ribbon region. The Rib43a–tubulin interaction leads to an elongated tubulin dimer distance every 2 dimers. In addition, the tubulin lattice structure with missing microtubule inner proteins (MIPs) by sarkosyl treatment shows significant longitudinal compaction and lateral angle change between protofilaments. These results are evidence that the MIPs directly affect and stabilize the tubulin lattice. It suggests that the doublet microtubule is an intrinsically stressed filament and that this stress could be manipulated in the regulation of ciliary waveforms