Phenotypic heterogeneity associated with KIF21A: Two new cases and review of the literature.

Our understanding of genetic and phenotypic heterogeneity associated with the clinical spectrum of rare diseases continues to expand. Thorough phenotypic descriptions and model organism functional studies are valuable tools in dissecting the biology of the disease process. Kinesin genes are well known to be associated with specific disease phenotypes and a subset of kinesin genes, including KIF21A, have been associated with more than one disease. Here we report two patients with KIF21A variants identified by exome sequencing; one with biallelic variants, supporting a novel KIF21A related syndrome with recessive inheritance and the second report of this condition, and another with a heterozygous de novo variant allele representing a phenotypic expansion of the condition described to date. We provide detailed phenotypic information on both families, including a novel neuropathology finding of neuroaxonal dystrophy associated with biallelic variants in KIF21A. Additionally, we studied the dominant variant in Saccharomyces cerevisiae to assess variant pathogenicity and found that this variant appears to impair protein function. KIF21A associated disease has mounting evidence for phenotypic heterogeneity; further patients and study of an allelic series are required to define the phenotypic spectrum and further explore the molecular etiology for each of these conditions.

Phenotype, genotype, and management of congenital fibrosis of extraocular muscles type 1 in 16 Chinese families.

PURPOSE: Congenital fibrosis of extraocular muscles type 1 (CFEOM1), a classical subtype of CFEOM, is characterized by restrictive ophthalmoplegia and ptosis. It is mainly caused by aberrant neural innervation of the extraocular muscles. This study aimed to investigate the genetic characteristics and clinical manifestations of CFEOM1 in Chinese families. METHODS: The clinical data, including ocular examinations, magnetic resonance imaging (MRI), and surgical procedures of affected individuals from 16 Chinese CFEOM1 families, were collected. The genomic DNA of 16 probands and their family members were sequenced for causative KIF21A gene mutations. Linkage analysis using microsatellite markers across KIF21A was also conducted. RESULTS: Affected individuals were presented with bilateral non-progressive ptosis, restricted horizontal eye movement, fixed infraduction of both eyes, compensatory chin-up head position, and neuromuscular abnormalities. Three heterozygous KIF21A mutations, c.2860C > T (p.R954W) (in eight families), c.2861G > T (p.R954L) (in two families), and c.2861G > A (p.R954Q) (in two families) were identified, which implied that hotspot mutations were common in Chinese CFEOM1 families. Germline Mosaicism was likely to be the cause of affected individuals with asymptomatic parents without KIF21A mutations presented in the eight families. Two affected individuals underwent modified levator muscle complex suspension surgery and achieved a good result without any complications. CONCLUSION: Instead of evaluating the whole CFEOM1 gene variant, hotspot mutations could be given priority for screening. The occurrence of germline mosaicism has to be taken into account in genetic counseling. Patients with CFEOM1 who have ptosis may benefit from an innovative surgical procedure called modified levator muscle complex suspension.

KIF21A regulates breast cancer aggressiveness and is prognostic of patient survival and tumor recurrence.

PURPOSE: Invasion of carcinoma cells into surrounding tissue affects breast cancer staging, influences choice of treatment, and impacts on patient outcome. KIF21A is a member of the kinesin superfamily that has been well-studied in congenital extraocular muscle fibrosis. However, its biological relevance in breast cancer is unknown. This study investigated the functional roles of KIF21A in this malignancy and examined its expression pattern in breast cancer tissue. METHODS: The function of KIF21A in breast carcinoma was studied in vitro by silencing its expression in breast cancer cells and examining the changes in cellular activities. Immunohistochemical staining of breast cancer tissue microarrays was performed to determine the expression patterns of KIF21A. RESULTS: Knocking down the expression of KIF21A using siRNA in MDA-MB-231 and MCF7 human breast cancer cells resulted in significant decreases in tumor cell migration and invasiveness. This was associated with reduced Patched 1 expression and F-actin microfilaments. Additionally, the number of focal adhesion kinase- and paxillin-associated focal adhesions was increased. Immunohistochemical staining of breast cancer tissue microarrays showed that KIF21A was expressed in both the cytoplasmic and nuclear compartments of carcinoma cells. Predominance of cytoplasmic KIF21A was significantly associated with larger tumors and high grade cancer, and prognostic of cause-specific overall patient survival and breast cancer recurrence. CONCLUSION: The data demonstrates that KIF21A is an important regulator of breast cancer aggressiveness and may be useful in refining prognostication of this malignant disease.

Structural analyses of key features in the KANK1·KIF21A complex yield mechanistic insights into the cross-talk between microtubules and the cell cortex.

The cross-talk between dynamic microtubules and the cell cortex plays important roles in cell division, polarity, and migration. A critical adaptor that links the plus ends of microtubules with the cell cortex is the KANK N-terminal motif and ankyrin repeat domains 1 (KANK1)/kinesin family member 21A (KIF21A) complex. Genetic defects in these two proteins are associated with various cancers and developmental diseases, such as congenital fibrosis of the extraocular muscles type 1. However, the molecular mechanism governing the KANK1/KIF21A interaction and the role of the conserved ankyrin (ANK) repeats in this interaction are still unclear. In this study, we present the crystal structure of the KANK1·KIF21A complex at 2.1 Å resolution. The structure, together with biochemical studies, revealed that a five-helix-bundle-capping domain immediately preceding the ANK repeats of KANK1 forms a structural and functional supramodule with its ANK repeats in binding to an evolutionarily conserved peptide located in the middle of KIF21A. We also show that several missense mutations present in cancer patients are located at the interface of the KANK1·KIF21A complex and destabilize its formation. In conclusion, our study elucidates the molecular basis underlying the KANK1/KIF21A interaction and also provides possible mechanistic explanations for the diseases caused by mutations in KANK1 and KIF21A.

Genetic investigation in a four-generation Chinese family with congenital fibrosis of extraocular muscles and keratoconus.

Here, we have reported the genetic and clinical characteristics of four generations of a family patient from China with congenital fibrosis of extraocular muscles 1 (CFEOM1) and keratoconus (KC). The history of diseases, clinical observations, and blood samples of all family members were collected. A total of 100 healthy participants were recruited as normal controls. The whole exome sequencing of the genomic DNA and polymerase chain reaction were performed on samples obtained from the controls and their family members to verify the gene variants. The functional analyses of the variants were performed by using different software. Two single nucleotide polymorphisms were detected in the proband and other patients in his families, including a heterozygous missense variation, g.39726207C > T (c.2860C > T, p.R954W, rs121912585), in the third highly conserved coiled-coil domain of KIF21A, and a heterozygous missense variant, g.30664732A > C (c.136A > C, p.S46R, rs200111443) in TGFBR2. The variant p.R954W in KIF21A was predicted to be pathogenic using software, whereas p.S46R in TGFBR2 was predicted to be of uncertain significance (VUS). Thus, KC might have occurred in the proband and his daughter because of a combination of genetic mutations and involuntary eye rubbing induced by CFEOM1. This is the first case of concomitant KC in a family having CFEOM1. Thus, the study provides new information about patients with KC having CFEOM1. Furthermore, the study suggests that attention should be paid to the early detection and diagnosis of KC in patients with CFEOM1.

TUBB3 and KIF21A in neurodevelopment and disease.

Neuronal migration and axon growth and guidance require precise control of microtubule dynamics and microtubule-based cargo transport. TUBB3 encodes the neuronal-specific ß-tubulin isotype III, TUBB3, a component of neuronal microtubules expressed throughout the life of central and peripheral neurons. Human pathogenic TUBB3 missense variants result in altered TUBB3 function and cause errors either in the growth and guidance of cranial and, to a lesser extent, central axons, or in cortical neuronal migration and organization, and rarely in both. Moreover, human pathogenic missense variants in KIF21A, which encodes an anterograde kinesin motor protein that interacts directly with microtubules, alter KIF21A function and cause errors in cranial axon growth and guidance that can phenocopy TUBB3 variants. Here, we review reported TUBB3 and KIF21A variants, resulting phenotypes, and corresponding functional studies of both wildtype and mutant proteins. We summarize the evidence that, in vitro and in mouse models, loss-of-function and missense variants can alter microtubule dynamics and microtubule-kinesin interactions. Lastly, we highlight additional studies that might contribute to our understanding of the relationship between specific tubulin isotypes and specific kinesin motor proteins in health and disease.

Structural model of microtubule dynamics inhibition by kinesin-4 from the crystal structure of KLP-12 -tubulin complex.

Kinesin superfamily proteins are microtubule-based molecular motors driven by the energy of ATP hydrolysis. Among them, the kinesin-4 family is a unique motor that inhibits microtubule dynamics. Although mutations of kinesin-4 cause several diseases, its molecular mechanism is unclear because of the difficulty of visualizing the high-resolution structure of kinesin-4 working at the microtubule plus-end. Here, we report that KLP-12, a C. elegans kinesin-4 ortholog of KIF21A and KIF21B, is essential for proper length control of C. elegans axons, and its motor domain represses microtubule polymerization in vitro. The crystal structure of the KLP-12 motor domain complexed with tubulin, which represents the high-resolution structural snapshot of the inhibition state of microtubule-end dynamics, revealed the bending effect of KLP-12 for tubulin. Comparison with the KIF5B-tubulin and KIF2C-tubulin complexes, which represent the elongation and shrinking forms of microtubule ends, respectively, showed the curvature of tubulin introduced by KLP-12 is in between them. Taken together, KLP-12 controls the proper length of axons by modulating the curvature of the microtubule ends to inhibit the microtubule dynamics.

From meter-long structures that allow nerve cells to stretch across a body to miniscule 'hairs' required for lung cells to clear mucus, many life processes rely on cells sporting projections which have the right size for their role. Networks of hollow filaments known as microtubules shape these structures and ensure that they have the appropriate dimensions. Controlling the length of microtubules is therefore essential for organisms, yet how this process takes place is still not fully elucidated. Previous research has shown that microtubules continue to grow when their end is straight but stop when it is curved. A family of molecular motors known as kinesin-4 participate in this process, but the exact mechanisms at play remain unclear. To investigate, Tuguchi, Nakano, Imasaki et al. focused on the KLP-12 protein, a kinesin-4 equivalent which helps to controls the length of microtubules in the tiny worm Caenorhabditis elegans. They performed genetic manipulations and imaged the interactions between KLP-12 and the growing end of a microtubule using X-ray crystallography. This revealed that KLP-12 controls the length of neurons by inhibiting microtubule growth. It does so by modulating the curvature of the growing end of the filament to suppress its extension. A 'snapshot' of KLP-12 binding to a microtubule at the resolution of the atom revealed exactly how the protein helps to bend the end of the filament to prevent it from growing further. These results will help to understand how nerve cells are shaped. This may also provide insights into the molecular mechanisms for various neurodegenerative disorders caused by problems with the human equivalents of KLP-12, potentially leading to new therapies.

KIF21A pathogenic variants cause congenital fibrosis of extraocular muscles type 3.

Background: Congenital fibrosis of the extraocular muscles (CFEOM) is characterized by ptosis and non-progressive restrictive ophthalmoplegia. CFEOM1 is a stereotypical phenotype with isolated bilateral ptosis, bilateral ophthalmoplegia, absent upgaze, and globe infraduction. CFEOM3 is a more variable phenotype that can include unilateral disease, absent ptosis, residual upgaze, and/or orthotropia. Most cases of CFEOM1 result from recurrent heterozygous KIF21A missense mutations and less commonly from recurrent heterozygous TUBB3 missense mutations. While most cases of CFEOM3 result from recurrent heterozygous TUBB3 missense mutations, several pedigrees harbored pathogenic variants in KIF21A. Here, we asked if Lebanese pedigrees with CFEOM3 harbor pathogenic variants in TUBB3 or KIF21A.Materials and Methods: Families affected with congenital cranial dysinnervation disorders were prospectively recruited from the American University of Beirut pediatric ophthalmology clinic and included two probands with CFEOM. KIF21A hotspot exons and TUBB3 coding sequence were sequenced. Available family members were sequenced for co-segregation analysis.Results: Both families were found to have CFEOM3 and to harbor pathogenic variants in KIF21A(OMIM 608283). A simplex proband with CFEOM3 from a consanguineous Iraqi family harbored a de novo heterozygous KIF21A c.2860 C > T variant (p.R954W); this variant accounts for the majority of reported KIF21A mutations but is typically implicated in CFEOM1. A Lebanese father with CFEOM3 and his son with CFEOM1 segregated a heterozygous KIF21A c.2830 G > C variant (p.E944Q), previously reported in an individual with CFEOM1.Conclusions: These results support prior reports of KIF21A mutations as a rare cause of CFEOM3. These families are Middle Eastern or Chinese, supporting a genetic modifier in these populations.

A rare case of congenital fibrosis of extraocular muscle type 1A due to KIF21A mutation with Marcus Gunn jaw-winking phenomenon.

BACKGROUND: Congenital fibrosis of the extraocular muscles (CFEOM1) is classically a congenital, non-progressive, restrictive strabismus syndrome characterized by bilateral ptosis and ophthalmoplegia with an infraducted position of the globes. This autosomal dominant syndrome is caused by mutations in the KIF21A gene. METHODS AND RESULTS: In this report we describe a 5-year-old boy, and his mother, both of whom have a mutation in the KIF21A gene, who possesses typical features of CFEOM1 syndrome. Besides displaying typical features of CFEOM1, he demonstrated Marcus Gunn jaw-winking phenomenon. The patient additionally had a positive family history of such features. CONCLUSION: This is first report of the coexistence of CFEOM and Marcus Gunn jaw-winking phenomenon in a patient with a KIF21A mutation from Turkey. We explain the phenotypic findings associated with mutations in KIF21A including CFEOM1A and Marcus Gunn jaw-winking phenomenon.