Primary Ciliary Dyskinesia Caused by Homozygous Mutation in DNAL1, Encoding Dynein Light Chain 1

Abstract

In primary ciliary dyskinesia (PCD), genetic defects affecting motility of cilia and flagella cause chronic destructive airway disease, randomization of left-right body asymmetry, and, frequently, male infertility. The most frequent defects involve outer and inner dynein arms (ODAs and IDAs) that are large multiprotein complexes responsible for cilia-beat generation and regulation, respectively. Although it has long been suspected that mutations in DNAL1 encoding the ODA light chain1 might cause PCD such mutations were not found. We demonstrate here that a homozygous point mutation in this gene is associated with PCD with absent or markedly shortened ODA. The mutation (NM_031427.3: c.449A>G; p.Asn150Ser) changes the Asn at position150, which is critical for the proper tight turn between the β strand and the α helix of the leucine-rich repeat in the hydrophobic face that connects to the dynein heavy chain. The mutation reduces the stability of the axonemal dynein light chain 1 and damages its interactions with dynein heavy chain and with tubulin. This study adds another important component to understanding the types of mutations that cause PCD and provides clinical information regarding a specific mutation in a gene not yet known to be associated with PCD.

Functional Architecture of the Outer Arm Dynein Conformational Switch

“…Pro residues can exist in either cis or trans conformations (Fig. 7), and molecular modeling revealed that a cis-Pro residue is readily accommodated within the wild-type structure 15 N HSQC spectra for wild-type and the M182A, D185G, D185P, R189E, and R196A mutant forms of LC1 are shown in the lower panels. Each peak represents an amide resonance derived from the peptide backbone, Asn/Gln side chains, or the Trp indole ring.…”

“…Even so, the number of residues exhibiting significant ⌬␦ values suggests that the observed phenotypes cannot necessarily be ascribed to the mutation of a particular residue but rather may reflect a more global perturbation or reorientation of this domain. To address this directly, we reassigned the backbone for 13 C, 15 Nlabeled D185G LC1 from 1 H-15 N HSQC, HNCA, HNCO, and HBHA(CO)NH spectra and performed a TALOSϩ backbone chemical shift analysis. This revealed that the overall helical structure of the C-terminal domain was not greatly perturbed but that the and angles for residues surrounding the mutation site were significantly changed (Fig.…”

“…In humans, a homozygous mutation within the last LRR of the LC1 orthologue DNAL1 has recently been demonstrated to cause primary ciliary dyskinesia (15). This LC consists of an N-terminal helix followed by six ␤␤␣ motifs derived from the LRRs and a C-terminal helical domain that protrudes from the main protein axis defined by the LRR barrel (16).…”

“…To obtain the complete backbone assignments for the D185G form of LC1, the 1 H-15 N HSQC, HNCA, HNCO, and HBHA(CO)NH spectra were obtained from a 13 C, 15 N-labeled sample using a Varian 500-MHz NMR spectrometer equipped with a cryogenic probe; the HNCACB experiment did not yield a usable spectrum due to the large size of LC1. Peak assignments were made with XEASY, and the backbone torsion angles were predicted from the chemical shifts using TALOSϩ (22 15 N HSQC spectrum from 13 C, 15 N-labeled R189E LC1 was sufficiently poorly resolved that it was not feasible to reassign the entire backbone for this mutant form. Circular Dichroism Spectroscopy-The concentrations of purified fully denatured wild-type, D185G, and N150S LC1 proteins in 2.5 mM Tris⅐Cl, pH 6.8, 50 mM NaCl were determined by absorbance at 280 nm using an extinction coefficient (⑀) of 17,810 liters⅐mol Ϫ1 ⅐cm Ϫ1 .…”

Diagnostic Methods in Primary Ciliary Dyskinesia

“…To date mutations have been identified in 6 genes encoding for proteins that are part of the ODA (DNAH5, DNAI1, DNAI2, DNAL1, NME8 (TXNDC3) and DNAH11) [12,[71][72][73][74][75]. Mutations in DNAH5 and DNAI1 are thought to account for the largest proportion of PCD patients: 30% and 9% respectively [19,[76][77][78][79].…”