Phenotypic Mutation 'dwarf' (pdf version)
Alleledwarf
Mutation Type missense
Chromosome12
Coordinate111,762,037 bp (GRCm39)
Base Change T ⇒ C (forward strand)
Gene Klc1
Gene Name kinesin light chain 1
Synonym(s) Kns2
Chromosomal Location 111,725,283-111,774,278 bp (+) (GRCm39)
MGI Phenotype FUNCTION: Conventional kinesin is a tetrameric molecule composed of two heavy chains and two light chains, and transports various cargos along microtubules toward their plus ends. The heavy chains provide the motor activity, while the light chains bind to various cargos. This gene encodes a member of the kinesin light chain family. It associates with kinesin heavy chain through an N-terminal domain, and six tetratricopeptide repeat (TPR) motifs are thought to be involved in binding of cargos such as vesicles, mitochondria, and the Golgi complex. Thus, kinesin light chains function as adapter molecules and not motors per se. Although previously named "kinesin 2", this gene is not a member of the kinesin-2 / kinesin heavy chain subfamily of kinesin motor proteins. Extensive alternative splicing produces isoforms with different C-termini that are proposed to bind to different cargos; however, the full-length nature of some of these variants has not been determined. [provided by RefSeq, Jul 2008]
PHENOTYPE: Mice homozygous for disruptions in this gene are significantly smaller than normal. [provided by MGI curators]
Accession Number

NCBI RefSeq: NM_008450 (variant a), NM_001025358 (variant b), NM_001025359 (variant c), NM_001025360 (variant d), NM_001025361 (variant e), NM_001025363 (variant f), NM_001025362 (variant g), NM_001081959 (variant h); MGI:107978

MappedYes 
Amino Acid Change Isoleucine changed to Threonine
Institutional SourceBeutler Lab
Gene Model predicted gene model for protein(s): [ENSMUSP00000082004] [ENSMUSP00000113171] [ENSMUSP00000113237] [ENSMUSP00000113997] [ENSMUSP00000120491]
AlphaFold no structure available at present
SMART Domains Protein: ENSMUSP00000082004
Gene: ENSMUSG00000021288
AA Change: I569T

DomainStartEndE-ValueType
coiled coil region 86 156 N/A INTRINSIC
low complexity region 158 179 N/A INTRINSIC
low complexity region 188 206 N/A INTRINSIC
Pfam:TPR_10 212 253 3.1e-9 PFAM
TPR 255 288 3.81e-1 SMART
TPR 297 330 1.16e-5 SMART
TPR 339 372 4.77e-2 SMART
TPR 381 414 2.78e-3 SMART
TPR 464 497 4.93e1 SMART
Predicted Effect probably damaging

PolyPhen 2 Score 0.997 (Sensitivity: 0.41; Specificity: 0.98)
(Using ENSMUST00000084941)
SMART Domains Protein: ENSMUSP00000113171
Gene: ENSMUSG00000021288

DomainStartEndE-ValueType
Pfam:Rab5-bind 80 254 8.3e-69 PFAM
Pfam:TPR_10 212 253 7.2e-9 PFAM
TPR 255 288 3.81e-1 SMART
TPR 297 330 1.16e-5 SMART
TPR 339 372 4.77e-2 SMART
TPR 381 414 2.78e-3 SMART
TPR 464 497 4.93e1 SMART
Predicted Effect probably benign
SMART Domains Protein: ENSMUSP00000113237
Gene: ENSMUSG00000021288

DomainStartEndE-ValueType
coiled coil region 86 156 N/A INTRINSIC
low complexity region 158 179 N/A INTRINSIC
low complexity region 188 206 N/A INTRINSIC
Pfam:TPR_10 212 253 3.2e-9 PFAM
TPR 255 288 3.81e-1 SMART
TPR 297 330 1.16e-5 SMART
TPR 339 372 4.77e-2 SMART
TPR 381 414 2.78e-3 SMART
TPR 464 497 4.93e1 SMART
Predicted Effect probably benign
SMART Domains Protein: ENSMUSP00000113997
Gene: ENSMUSG00000021288
AA Change: I561T

DomainStartEndE-ValueType
Pfam:Rab5-bind 80 254 1e-68 PFAM
Pfam:TPR_10 212 253 8.4e-9 PFAM
TPR 255 288 3.81e-1 SMART
TPR 297 330 1.16e-5 SMART
TPR 339 372 4.77e-2 SMART
TPR 381 414 2.78e-3 SMART
TPR 464 497 2.99e1 SMART
Predicted Effect probably damaging

PolyPhen 2 Score 0.994 (Sensitivity: 0.69; Specificity: 0.97)
(Using ENSMUST00000122300)
SMART Domains Protein: ENSMUSP00000120491
Gene: ENSMUSG00000021288

DomainStartEndE-ValueType
Pfam:TPR_1 1 25 1.9e-4 PFAM
Pfam:TPR_7 1 36 1.9e-4 PFAM
Pfam:TPR_10 75 112 7.8e-9 PFAM
Pfam:TPR_1 77 98 1.4e-4 PFAM
Pfam:TPR_7 78 129 1.7e-5 PFAM
Predicted Effect probably benign
Meta Mutation Damage Score 0.7446 question?
Is this an essential gene? Possibly nonessential (E-score: 0.498) question?
Phenotypic Category Autosomal Recessive
Candidate Explorer Status loading ...
Single pedigree
Linkage Analysis Data
Penetrance  
Alleles Listed at MGI

All Mutations and Alleles(55) : Gene trapped(48) Targeted(5) Transgenic(2)

Lab Alleles
AlleleSourceChrCoordTypePredicted EffectPPH Score
IGL00594:Klc1 APN 12 111743318 missense probably damaging 1.00
IGL00940:Klc1 APN 12 111753932 missense probably damaging 1.00
IGL02206:Klc1 APN 12 111744550 unclassified probably benign
IGL02487:Klc1 APN 12 111738886 missense probably damaging 0.99
IGL02490:Klc1 APN 12 111748210 missense possibly damaging 0.89
IGL02830:Klc1 APN 12 111743341 missense probably damaging 0.99
IGL03121:Klc1 APN 12 111748076 unclassified probably benign
IGL03253:Klc1 APN 12 111748078 unclassified probably benign
IGL03376:Klc1 APN 12 111742387 missense probably damaging 0.97
F5770:Klc1 UTSW 12 111741006 missense probably benign 0.09
R0031:Klc1 UTSW 12 111743467 missense probably damaging 0.99
R0239:Klc1 UTSW 12 111751758 splice site probably benign
R1647:Klc1 UTSW 12 111743321 missense probably damaging 1.00
R1648:Klc1 UTSW 12 111743321 missense probably damaging 1.00
R1892:Klc1 UTSW 12 111748261 critical splice donor site probably null
R2940:Klc1 UTSW 12 111772451 missense possibly damaging 0.49
R4829:Klc1 UTSW 12 111762037 missense probably damaging 1.00
R4849:Klc1 UTSW 12 111748129 missense probably damaging 1.00
R5309:Klc1 UTSW 12 111762055 missense possibly damaging 0.82
R5312:Klc1 UTSW 12 111762055 missense possibly damaging 0.82
R5637:Klc1 UTSW 12 111740842 missense probably damaging 1.00
R5706:Klc1 UTSW 12 111762061 missense possibly damaging 0.65
R6623:Klc1 UTSW 12 111772475 missense probably damaging 1.00
R6920:Klc1 UTSW 12 111754019 missense probably damaging 1.00
R7109:Klc1 UTSW 12 111743299 missense probably benign 0.22
R7538:Klc1 UTSW 12 111751879 missense probably benign 0.01
R8051:Klc1 UTSW 12 111748384 missense possibly damaging 0.58
R8719:Klc1 UTSW 12 111772509 critical splice donor site probably benign
R8995:Klc1 UTSW 12 111743344 missense probably damaging 1.00
R9420:Klc1 UTSW 12 111738950 missense probably damaging 0.99
V7580:Klc1 UTSW 12 111741006 missense probably benign 0.09
V7581:Klc1 UTSW 12 111741006 missense probably benign 0.09
Mode of Inheritance Autosomal Recessive
Local Stock
Repository
Last Updated 2019-09-04 9:41 PM by Anne Murray
Record Created 2016-12-24 4:03 PM
Record Posted 2017-03-31
Phenotypic Description

Figure 1. Dwarf mice exhibited reduced body weights compared to wild-type controls. Scaled weight data are shown. Abbreviations: WT, wild-type; REF, homozygous reference mice; HET, heterozygous variant mice; VAR, homozygous variant mice. Mean (μ) and standard deviation (σ) are indicated.

The dwarf phenotype was identified among N-ethyl-N-nitrosourea (ENU)-mutagenized G3 mice of the pedigree R4829, some of which showed reduced body weights compared to wild-type littermate controls (Figure 1).

Nature of Mutation

Figure 2. Linkage mapping of the reduced body weights using a recessive model of inheritance. Manhattan plot shows -log10 P values (Y-axis) plotted against the chromosome positions of 101 mutations (X-axis) identified in the G1 male of pedigree R4829. Weight data are shown for single locus linkage analysis without consideration of G2 dam identity. Horizontal pink and red lines represent thresholds of P = 0.05, and the threshold for P = 0.05 after applying Bonferroni correction, respectively.

Whole exome HiSeq sequencing of the G1 grandsire identified 101 mutations. The body weight phenotype was linked to two genes on chromosome 12: Klc1 and Adssl1. The mutation in Klc1 was presumed to be causative because the dwarf weight phenotype mimics other known alleles of Klc1 (see MGI for a list of Klc1 alleles). The Kcl1 mutation is a T to C transition at base pair 111,795,603 (v38) on chromosome 12, or base pair 36,755 in the GenBank genomic region NC_000078 for the Kcl1 gene.  Linkage was found with a recessive model of inheritance, wherein three variant homozygotes departed phenotypically from 14 homozygous reference mice and 35 heterozygous mice with a P value of 4.234 x 10-7 (Figure 2).  

The mutation corresponds to residue 1,901 in the mRNA sequence NM_001025360 within exon 14 of 16 total exons.

 
1885 AAACTCCGGGCTTCCATTAGACGCAGCAGTGAG

564  -K--L--R--A--S--I--R--R--S--S--E- (NP_001020531 & NP_001020532)

 

The mutated nucleotide is indicated in red.  The mutation results in an isoleucine (I) to threonine (T) substitution at position 569 (I569T) in the kinesin light chain 1 (variants d and e) protein and I561T in kinesin light chain 1 (variant c), and is strongly predicted by PolyPhen-2 to be damaging (score = 0.997).

Illustration of Mutations in
Gene & Protein
Protein Prediction
Figure 3. Domain organization of mouse KLC1. There are two major domains in KLC1: an N-terminal coiled-coil (heptad repeat) domain and six imperfect repeats of a 34 amino acid tetra-trico peptide repeat (TPR) domain at the C-terminus. The location of the dwarf mutation is indicated. 

Klc1 encodes KLC1, a light chain of kinesin-1.  There are four kinesin light chains in humans: KLC1, KLC2, KLC3, and KLC4. There are two major domains in KLC1: an N-terminal coiled-coil (heptad repeat) domain and six imperfect repeats of a 34 amino acid tetra-trico peptide repeat (TPR) domain at the C-terminus (Figure 3(1;2). The highly conserved N-terminal coiled-coil domain mediates binding to kinesin heavy chains (3). TPR domains often function in protein-protein interactions. For example, mitochondrial fission protein dynamin 1-like protein (Dnm1L) interacts with the TPR domain of KLC1 (4). The association between Dnm1L and KLC1 indicates that KLC1 may have a role in post-fission mitochondrial transport. KLC1 also interacts with JNK-interacting protein 1 (JIP1) and JIP3 via the TRP domain (2). Residues in the third TPR domain facilitate cargo binding. The inter-TPR loops between the second and third TPR domains is a third cargo-binding site (2). Additional binding partners of KLC1 include Huntingtin-associated protein-1 (HAP1), alcadein-1 (ALC1), torsin A, collapsing response mediator protein-1 (CRMP1), KIDINS220, and Daxx [reviewed in (2)]. The KLC1 TPR region has 13 α-helices; 12 of the α-helices are TPR1 through TPR6, and one non-TPR helix occurs between TPR5 and TPR6 (2). Each TPR domain has a helix-turn-helix arrangement and multiple TPR domains arrange in a superhelical conformation (2).

KLC1 phosphorylation regulates kinesin activity (5;6). KLC1 has a consensus AMPK phosphorylation site at Ser520. Ser460 is a putative MAPK target site; ERK phosphorylates the site in vitro (7). Mutation of Ser460 to an alanine (S460A) results in increased binding of KLC1 to calsyntenin-1 (7). Mutation of Ser460 to aspartate (S460D) (to mimic permanent phosphorylation) reduced KLC1 binding to calsyntenin-1. Tumor necrosis factor activates two putative kinesin light chain kinases (PKA and a MAPK), which results in hyperphosphorylation of KLCs and impairs kinesin motor activity (8).

Nineteen KLC1 alternatively spliced variants have been identified in humans (9). There are at least seven putative Klc1 splice variants in the mouse (9-11). The protein products of the splice variants differ at the C-termini, which mediates different cargo binding (9). The KLC1B and 1C splice variants are associated with the ER and mitochondria, while the KLC1D and 1E are associated with the Golgi apparatus (10-12). The KLC1B isoform is essential for rough endoplasmic reticulum motility (12). KLC1C is required for transendothelial migration (13). KLC1E is a modifier of amyloid-β accumulation in mice (14). In Alzheimer’s disease patients, the levels of KLC1E were higher than that in unaffected individuals.

The dwarf mutation results in an isoleucine (I) to threonine (T) substitution at position 569 (I569T) in the kinesin light chain 1 protein (variants d and e) and I561T in kinesin light chain 1 protein (variant c). The affected residue is an undefined region at the C-terminus of KLC1.

Expression/Localization

KLC1 is highly expressed in neurons (15;16).

Background
Figure 4. Kinesins are molecular motors that transport cargo along microtubules in an ATP-dependent manner. 

Kinesins are molecular motors that transport organelles, mediate cellular trafficking between the Golgi apparatus and the ER, promote mitochondrial placement and lysosomal movement, and move chromosomes along microtubules during cell division in an ATP-dependent manner (Figure 4(16). Kinesins are tetrameric proteins containing two heavy chains (i.e., KIF5A, KIF5B, or KIF5C) and two light chains (i.e., KLC1, KLC2, KLC3, or KLC4). KIF5A and KIF5C are neuron-specific, while KLC1 is enriched in neurons. KIF5B and KLC2 are ubiquitously expressed; the expression pattern of KLC3 is unknown (17). The heavy chains mediate tubulin binding and contain ATPase domains, while the light chains are required for attachment of the organelle that will be moved and stabilize the inactive conformation of kinesin (18). There are over 30 different cargoes moved by kinesin 1, including synaptic vesicles, amyloid precursor protein, tau, neurofilaments, post-synaptic neurochemical receptors, phagosomes in the retinal pigment epithelium, Mn2+ in the optic nerve, Huntingtin-associated protein-1, insulin granules, mitochondria, and endoplasmic reticulum and Golgi membranes [reviewed in (19)].

Klc1-deficient (Klc1-/-) mice were smaller than wild-type mice and exhibited motor disabilities (16)). Heavy chain (KIF5A and KIF5B) activation and targeting were aberrant in the Klc1-/- mice. Additional studies found that, with age, the Klc1-/- mice exhibited axonopathies with cytoskeletal disorganization and aberrant cargo accumulation (i.e., a decrease in the proportion of particles moving in the anterograde direction and an increase in the number of retrograde particles) (20). Reductions in the white matter of the spinal cord and brain were observed in the older Klc1-/- mice (20). In addition, there was a reduction of large caliber axons in peripheral motor roots in the older Klc1-/- mice (20). In the Klc1-/- mice, the association of amyloid precursor protein (APP) with microtubules as well as the axonal transport of APP was reduced (17). With age, Klc1-/- mice exhibited retinal pigment epithelium pathogenesis with accumulation of RPE and sub-RPE deposits as well as oxidative and inflammatory stress responses (21).

Faulty axonal transport may contribute to neurodegenerative diseases in humans (22). In Alzheimer’s disease patients, the level of KLC1 expression was reduced in the frontal cortex (23). Morel et al. proposed that loss of KLC1 expression in the frontal cortex of Alzheimer’s disease patients as well as GSK-3β-mediated phosphorylation of KLC1 contribute to disturbances of axoplasmic flows and synaptic integrity in Alzheimer’s disease (23). KLC1 is a proposed susceptibility gene for age-related cataracts (24;25).

Putative Mechanism

Similar to the Klc1-/- mice, the dwarf mice are smaller in size than their wild-type littermates. Motor defects were not observed in the dwarf mice. The phenotype of the dwarf mice indicates some loss of KLC1dwarf function. How mutations in KLC1 lead to reduced body size is unknown.

Primers PCR Primer
dwarf_pcr_F: GTAACATGTCTGTGCTCACTGC
dwarf_pcr_R: TGAACAGAATCAAGGCGCTC

Sequencing Primer
dwarf_seq_F: TCACTGCAGGCTGCTGG
dwarf_seq_R: TTTGAGCCTCGACCAGCCAG
Genotyping

PCR program

1) 94°C 2:00
2) 94°C 0:30
3) 55°C 0:30
4) 72°C 1:00
5) repeat steps (2-4) 40x
6) 72°C 10:00
7) 4°C hold


The following sequence of 486 nucleotides is amplified (chromosome 12, + strand):


1   gtaacatgtc tgtgctcact gcaggctgct ggctctggca ggactcacgt gccagccagt
61  gcctggcagc agttggggtt gttgggctga gcatggggca ggaggggaca cccacagcgt
121 tacacttgtt cacgtgtttt tcaggatggc actggatctt taaagcgcag tggctccttt
181 agcaaactcc gggcttccat tagacgcagc agtgagaagc tggttaggaa gctgaaggga
241 ggaagctcac gggacagtga gccgaggaac cctgggtaac tatctcccga gcgcgcccct
301 tcccgccagt gttggttccg agccgcagcc catccgaggc agccttcctg gaactcaggg
361 ttggtcccag gagcggtcgg tcggtcgggt gctccgcctg ctgctttcca gggcaggttg
421 aggagctgtt gggaagctgg ctggtcgagg ctcaaaggga gcggtggagc gccttgattc
481 tgttca


Primer binding sites are underlined and the sequencing primers are highlighted; the mutated nucleotide is shown in red.

References
Science Writers Anne Murray
Illustrators Katherine Timer
AuthorsEmre Turer and Bruce Beutler