Strongly Truncated Dnaaf4 Plays a Conserved Role in Drosophila Ciliary Dynein Assembly as Part of an R2TP-Like Co-Chaperone Complex With Dnaaf6

Abstract

Axonemal dynein motors are large multi-subunit complexes that drive ciliary movement. Cytoplasmic assembly of these motor complexes involves several co-chaperones, some of which are related to the R2TP co-chaperone complex. Mutations of these genes in humans cause the motile ciliopathy, Primary Ciliary Dyskinesia (PCD), but their different roles are not completely known. Two such dynein (axonemal) assembly factors (DNAAFs) that are thought to function together in an R2TP-like complex are DNAAF4 (DYX1C1) and DNAAF6 (PIH1D3). Here we investigate the Drosophila homologues, CG14921/Dnaaf4 and CG5048/Dnaaf6. Surprisingly, Drosophila Dnaaf4 is truncated such that it completely lacks a TPR domain, which in human DNAAF4 is likely required to recruit HSP90. Despite this, we provide evidence that Drosophila Dnaaf4 and Dnaaf6 proteins can associate in an R2TP-like complex that has a conserved role in dynein assembly. Both are specifically expressed and required during the development of the two Drosophila cell types with motile cilia: mechanosensory chordotonal neurons and sperm. Flies that lack Dnaaf4 or Dnaaf6 genes are viable but with impaired chordotonal neuron function and lack motile sperm. We provide molecular evidence that Dnaaf4 and Dnaaf6 are required for assembly of outer dynein arms (ODAs) and a subset of inner dynein arms (IDAs).

Upregulation of ENKD1 disrupts cellular homeostasis to promote lymphoma development

“…PTGIS and EARS2 were demonstrated to regulate colon cancer metastasis (Cui et al, 2021; Sun et al, 2012). In addition, UBXN11, MALSU1, and PIH1D3 play important roles in the formation of actin stress fibers (Yadav et al, 2021), maintenance of mitochondrial homeostasis (Rebelo‐Guiomar et al, 2022), and ciliary dynamin assembly (Lennon et al, 2022). These findings suggest that ENKD1 may be involved in cellular homeostatic regulation through its interactions with these genes, which may further contribute to the occurrence and development of DLBCL.…”

PIH1D3-knockout rats exhibit full ciliopathy features and dysfunctional pre-assembly and loading of dynein arms in motile cilia

“…A full depletion of PIH1D3 in the KO rats induced robust phenotypes, including communicating hydrocephalus. The hydrocephalus phenotype is correlated with the deletion of PIH1D3 from the epithelial cells of ependyma and choroid plexus in the KO rats, although hydrocephalus was not reported in patients carrying a pathogenic mutation in PIH1D3 (Olcese et al, 2017;Paff et al, 2017;Paff et al, 2018;Aprea et al, 2021;Lennon et al, 2022). The difference in phenotypic expression between humans and rats possibly results from the difference in compensatory mechanisms in different species.…”

“…We reported a novel rat model that was depleted of PIH1D3 expression and developed the cardinal features of PIH1D3-related diseases ( Olcese et al, 2017 ; Paff et al, 2017 ; Paff et al, 2018 ; Aprea et al, 2021 ; Lennon et al, 2022 ). PIH1D3-KO rats exhibited situs inversus, defects in spermatocyte survival and mucociliary clearance, and perinatal hydrocephalus.…”

“…Our data suggest that PIH1D3 is implicated in both the assembly and trafficking of ODA and IDA, providing a novel insight into PIH1D3 functions. Preservation of function in mutated PIH1D3 affects the severity of disease phenotypes in patients (Olcese et al, 2017;Paff et al, 2017;Paff et al, 2018;Aprea et al, 2021;Lennon et al, 2022). Our PIH1D3-KO rats were generated by introducing a frameshift in exon 5 and inducing subsequent frameshift within exon 6, leading to the nonsense-mediated decay of mutant PIH1D3 transcripts.…”