The outer dynein arm assembly factor CCDC103 forms molecular scaffolds through multiple self‐interaction sites

Abstract

CCDC103 is a small protein with unusual biophysical properties that is required for outer dynein arm assembly on ciliary axonemes. Mutations in both human and zebrafish CCDC103 proteins lead to primary ciliary dyskinesia. Previous studies revealed that this protein can oligomerize and appears to be arrayed along the entire length of the ciliary axoneme. CCDC103 also binds purified microtubules directly and indeed stabilizes them. Here we use biochemical approaches to identify two regions of CCDC103 that mediate self-interaction. In both cases, these associations are stable to heating in the presence of detergent and are not disrupted by strong reducing agents. One interaction region consists of a 27-residue inherently disorder segment that can mediate heat/detergent-resistant dimerization when attached to unrelated monomeric proteins. The second interface includes the C-terminal RPAP3_C alpha helical domain. Our data suggest that CCDC103 can form an unconventional polymer and we propose models for how the monomers might be organized. We also use molecular modeling of the RPAP3_C domain to determine the structural consequences of the pathogenic H154P mutation found in human PCD patients. K E Y W O R D S Axoneme, Chlamydomonas, cilia, dynein, flagella, microtubule ABBREVIATIONS: CCDC, coiled coil domain containing; FAP, flagella-associated protein; MBP, maltose-binding protein; ODA-DC, outer dynein arm docking complex; RPAP3, RNA polymerase II-associated protein 3; TCEP, Tris(2-carboxyethyl)phosphine.

Cilia-independent requirements for Ccdc103 promoting proliferation and migration of myeloid cells

“…Based on biochemical evidence showing that CCDC103 forms self-organizing oligomers, others have hypothesized that CCDC103 may function as a molecular scaffold, anchoring other proteins at the microtubule and facilitating their function (King and Patel-King, 2020). However, 10 very little is known about the network of CCDC103-interacting proteins.…”

“…Mutations in CCDC103 underlie approximately 4% of all PCD cases, but in certain geographic subpopulations where PCD is more prevalent it has been shown to be responsible for ∼20% of cases (Shoemark et al ., 2018). CCDC103 is a small 242 amino acid (AA) protein that consists of a central RPAP3_C domain flanked by N- and C-terminal coiled coils (King and Patel-King, 2020). It is critical for the proper docking and assembly of the outer dynein arms which facilitate ciliary motion, but it is also found localized throughout the cytoplasm of both ciliated and unciliated cells (Panizzi et al ., 2012).…”

“…It is critical for the proper docking and assembly of the outer dynein arms which facilitate ciliary motion, but it is also found localized throughout the cytoplasm of both ciliated and unciliated cells (Panizzi et al ., 2012). In vitro studies have shown that CCDC103 forms self-organizing oligomers, and that it binds periodically to cytoplasmic microtubules and can facilitate the stability of assembled microtubules (King and Patel-King, 2015; King and Patel-King, 2020). Despite these studies, if CCDC103 has cilia-independent requirements within the cytoplasm in vivo remains unknown.…”

Ccdc103 promotes myeloid cell proliferation and migration independent of motile cilia

“…Mutations in CCDC103 underlie ∼4% of all PCD cases, but in certain geographic subpopulations in which PCD is more prevalent it has been shown to be responsible for ∼20% of cases ( Shoemark et al, 2018 ). Human CCDC103 is a small 242-amino acid (AA) protein that consists of conserved central RPAP3_C and flanking N- and C-terminal coiled-coil domains ( King and Patel-King, 2020 ). CCDC103 is critical for the proper docking and assembly of the outer dynein arms, which facilitate ciliary motion, but is also found localized throughout the cytoplasm of both ciliated and non-ciliated cells ( Panizzi et al, 2012 ).…”

“…CCDC103 is critical for the proper docking and assembly of the outer dynein arms, which facilitate ciliary motion, but is also found localized throughout the cytoplasm of both ciliated and non-ciliated cells ( Panizzi et al, 2012 ). In vitro studies have shown that CCDC103 forms self-organizing oligomers, and that it binds periodically to cytoplasmic microtubules and can facilitate the stability of assembled microtubules ( King and Patel-King, 2015 , 2020 ). Despite these studies, it is not yet known whether CCDC103 has cilia-independent requirements in the cytoplasm in vivo.…”