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Destabilization of the IFT-B cilia core complex due to mutations in IFT81 causes a Spectrum of Short-Rib Polydactyly Syndrome

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ABSTRACT

Short-rib polydactyly syndromes (SRPS) and Asphyxiating thoracic dystrophy (ATD) or Jeune Syndrome are recessively inherited skeletal ciliopathies characterized by profound skeletal abnormalities and are frequently associated with polydactyly and multiorgan system involvement. SRPS are produced by mutations in genes that participate in the formation and function of primary cilia and usually result from disruption of retrograde intraflagellar (IFT) transport of the cilium. Herein we describe a new spectrum of SRPS caused by mutations in the gene IFT81, a key component of the IFT-B complex essential for anterograde transport. In mutant chondrocytes, the mutations led to low levels of IFT81 and mutant cells produced elongated cilia, had altered hedgehog signaling, had increased post-translation modification of tubulin, and showed evidence of destabilization of additional anterograde transport complex components. These findings demonstrate the importance of IFT81 in the skeleton, its role in the anterograde transport complex, and expand the number of loci associated with SRPS.

No MeSH data available.


IFT-B complex is destabilized in IFT81 mutants.(A) RNA levels of IFT81 in control and patient chondrocytes. β-Actin serves as loading control. (B) Protein levels of several components of IFT-B complex (IFT81, IFT74, IFT88 and IFT52), acetyl-tubulin and kinesin motor component (KIF3A). GADPH serves as a loading control. (C–H) Bar graphs showing statistical analyses (t-Test) for the replicates of each studied protein between control and R98-443. (I) Cartoon of the IFT-B complex transporting αβ-tubulin through a cilia microtubule.
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f2: IFT-B complex is destabilized in IFT81 mutants.(A) RNA levels of IFT81 in control and patient chondrocytes. β-Actin serves as loading control. (B) Protein levels of several components of IFT-B complex (IFT81, IFT74, IFT88 and IFT52), acetyl-tubulin and kinesin motor component (KIF3A). GADPH serves as a loading control. (C–H) Bar graphs showing statistical analyses (t-Test) for the replicates of each studied protein between control and R98-443. (I) Cartoon of the IFT-B complex transporting αβ-tubulin through a cilia microtubule.

Mentions: Cultured cells (chondrocytes) were only available for R98-443A and were used to assess the effect of the mutations on IFT81 mRNA and protein levels. RT-PCR demonstrated at least a 50 percent reduction in the transcript level in mutant cells relative to controls (Fig. 2A). Western blot analyses of IFT81 showed near complete loss of IFT81 in mutant cells (Fig. 2B). A non-specific band, that migrated slightly faster than IFT81, did not differ between control and mutant cells. These data demonstrate that compound heterozygosity for the mutations led to a significant loss of IFT81, more than could be accounted for by presence of one allele, suggesting that the missense mutation destabilized IFT81.


Destabilization of the IFT-B cilia core complex due to mutations in IFT81 causes a Spectrum of Short-Rib Polydactyly Syndrome
IFT-B complex is destabilized in IFT81 mutants.(A) RNA levels of IFT81 in control and patient chondrocytes. β-Actin serves as loading control. (B) Protein levels of several components of IFT-B complex (IFT81, IFT74, IFT88 and IFT52), acetyl-tubulin and kinesin motor component (KIF3A). GADPH serves as a loading control. (C–H) Bar graphs showing statistical analyses (t-Test) for the replicates of each studied protein between control and R98-443. (I) Cartoon of the IFT-B complex transporting αβ-tubulin through a cilia microtubule.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5035930&req=5

f2: IFT-B complex is destabilized in IFT81 mutants.(A) RNA levels of IFT81 in control and patient chondrocytes. β-Actin serves as loading control. (B) Protein levels of several components of IFT-B complex (IFT81, IFT74, IFT88 and IFT52), acetyl-tubulin and kinesin motor component (KIF3A). GADPH serves as a loading control. (C–H) Bar graphs showing statistical analyses (t-Test) for the replicates of each studied protein between control and R98-443. (I) Cartoon of the IFT-B complex transporting αβ-tubulin through a cilia microtubule.
Mentions: Cultured cells (chondrocytes) were only available for R98-443A and were used to assess the effect of the mutations on IFT81 mRNA and protein levels. RT-PCR demonstrated at least a 50 percent reduction in the transcript level in mutant cells relative to controls (Fig. 2A). Western blot analyses of IFT81 showed near complete loss of IFT81 in mutant cells (Fig. 2B). A non-specific band, that migrated slightly faster than IFT81, did not differ between control and mutant cells. These data demonstrate that compound heterozygosity for the mutations led to a significant loss of IFT81, more than could be accounted for by presence of one allele, suggesting that the missense mutation destabilized IFT81.

View Article: PubMed Central - PubMed

ABSTRACT

Short-rib polydactyly syndromes (SRPS) and Asphyxiating thoracic dystrophy (ATD) or Jeune Syndrome are recessively inherited skeletal ciliopathies characterized by profound skeletal abnormalities and are frequently associated with polydactyly and multiorgan system involvement. SRPS are produced by mutations in genes that participate in the formation and function of primary cilia and usually result from disruption of retrograde intraflagellar (IFT) transport of the cilium. Herein we describe a new spectrum of SRPS caused by mutations in the gene IFT81, a key component of the IFT-B complex essential for anterograde transport. In mutant chondrocytes, the mutations led to low levels of IFT81 and mutant cells produced elongated cilia, had altered hedgehog signaling, had increased post-translation modification of tubulin, and showed evidence of destabilization of additional anterograde transport complex components. These findings demonstrate the importance of IFT81 in the skeleton, its role in the anterograde transport complex, and expand the number of loci associated with SRPS.

No MeSH data available.