Crystal Structure of Bicc1 SAM Polymer and Mapping of Interactions between the Ciliopathy-Associated Proteins Bicc1, ANKS3, and ANKS6

Abstract

Head-to-tail polymers of sterile alpha motifs (SAM) can scaffold large macromolecular complexes. Several SAM-domain proteins that bind each other are mutated in patients with cystic kidneys or laterality defects, including the Ankyrin (ANK) and SAM domain-containing proteins ANKS6 and ANKS3, and the RNA-binding protein Bicc1. To address how their interactions are regulated, we first determined a high-resolution crystal structure of a Bicc1-SAM polymer, revealing a canonical SAM polymer with a high degree of flexibility in the subunit interface orientations. We further mapped interactions between full-length and distinct domains of Bicc1, ANKS3, and ANKS6. Neither ANKS3 nor ANKS6 alone formed macroscopic homopolymers in vivo. However, ANKS3 recruited ANKS6 to Bicc1, and the three proteins together cooperatively generated giant macromolecular complexes. Thus, the giant assemblies are shaped by SAM domains, their flanking sequences, and SAM-independent protein-protein and protein-mRNA interactions.

Role of Bicaudal C1 in renal gluconeogenesis and its novel interaction with the CTLH complex

“…However, co-transfection of full length ANKS3 or of the truncated mutant ANKS3ΔC lacking the C-terminal region distal of its SAM domain inhibited CTLH complex binding to HA-Bicc1 ( Fig 6E ), indicating that ANKS3 competes with CTLH complex for Bicc1. Previous structure analysis suggests that the SAM domain of ANKS3 interfaces with the ML and EH surfaces of the Bicc1 SAM domain, thereby allowing the bulky C-terminal region of ANKS3 to block Bicc1 polymer elongation [ 41 ]. To distinguish whether ML or EH surfaces of the Bicc1 SAM domain or their oligomerization also mediate CTLH complex binding, we mutated them individually [ 19 ] ( S3 Fig ).…”

Molecular and cellular basis of left–right asymmetry in vertebrates

“…However, it may be relevant that Bicc1, a putative RNA-binding protein specifically expressed at the node of mouse embryos, is essential for L-R asymmetry. 65 ) Furthermore, Bicc1 interacts with ANKS3 and ANKS6, 66 , 67 ) and ANKS3 mutations have been identified in patients with laterality defects. 68 ) Finally, Bicc1 also binds to Cerl2 mRNA in zebrafish and thereby inhibits its translation.…”

Nephronophthisis gene products display RNA-binding properties and are recruited to stress granules

“…We observed that NPH members interact with BICC1, the mammalian homologue of the Drosophila bicaudal C (BicC) 15 , and detailed structural analysis confirmed the interaction of BICC1 with ANKS3 and ANKS6 19 . Required during Drosophila oogenesis to down-regulate oskar mRNA at the anterior pole 20 , BicC contains three conserved N-terminal K-homology (KH) domains responsible for RNA binding, and a C-terminal Sterile Alpha Motif domain (SAM), which recruits BICC1 into RNA-processing bodies (P-bodies) 21 .…”

“…BICC1 affects the localization of ANKS3 and ANKS6 19 . In the absence of hippuristanol, BICC1 recruited both proteins into TIA-1-negative granules, likely representing RNA-processing bodies.…”

“…The presence of AGO2 mRNA in the BICC1 precipitate and the known localization of BICC1 to RNA-processing granules (P-bodies) led us to explore this interaction. Self-polymerization of the BICC1 by the SAM domain facilitates accumulation of BICC1 in cytoplasmic foci 19 , 29 . We confirmed that BICC1 accumulates in cytoplasmic granules.…”