Structural Analysis of Ciliogenesis Proteins Ttc29 and Lrrc56

Tynan Gardner

Ttc29 and Lrrc56 in Ciliogenesis

Cilia are widely conserved eukaryotic organelles that can serve two functions: sensing extracellular signals or beating back and forth to drive extracellular fluid flow. Mutations in genes involved in ciliogenesis lead to a variety of congenital diseases termed ciliopathies. Many proteins involved in assembling cilia are still being discovered and characterized, and uncovering their structures and protein-protein interactions will be vital to understanding the proper development and homeostasis of cilia. This project will focus on two proteins recently implicated in ciliogenesis both in laboratory and clinical settings.

Ttc29: Tetratricopeptide Repeat Protein 29

Mutations in Ttc29 have been shown to cause male subfertility with reduced sperm motility in both humans and mice. Recent work in the Wallingford and Marcotte labs has implicated the protein Ttc29 in inner and outer dynein arm transport to the cilia, which are required for motile cilia beating; morpholino knockdown of Ttc29 in Xenopus embryos prevents axonemal localization of ODAs and IDAs. Affinity Purification Mass Spectrometry has identified 4 candidate proteins that may interact with Ttc29.

Ttc29 localizes to foci (cyan) that are often adjacent to, but not a part of, DynAP organelles (magenta).

Summary of Ttc29 APMS Results. Line weight corresponds to significance of interaction.

Mutant Ttc29 allele from a patient with male infertility with asthenoteratospermia

Lrrc56: Leucine-Rich Repeat-Containing Protein 56

Lrrc56 is a poorly characterized protein that is required for ODA transport in algae and is implicated in intraflagellar transport in Trypanosomes, but has only recently been studied in vertebrates. Mutations in this gene have recently been reported to cause mucociliary clearance and laterality defects in clinical cases. The protein has been shown to localize to the DynAP, a liquid organelle that concentrates and organizes proteins required for Dynein assembly. Little is known about how the DynAP constituent proteins perform this function.

Mutant Lrrc56 alleles from three unrelated families with mucociliary clearance and laterality defects. Missense allele c.419T>C: p.(Leu140Pro)  was used for structural analysis.