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A novel mutation in β integrin reveals an integrin-mediated interaction between the extracellular matrix and cki-1/p27KIP1.

Kihira S, Yu EJ, Cunningham J, Cram EJ, Lee M - PLoS ONE (2012)

Bottom Line: RNAi of unc-52/perlecan, ina-1/α integrin, pat-4/ILK, and unc-97/PINCH resulted in abnormal CKI-1::GFP localization.These data also suggest that integrin plays a major role in maintaining proper CKI-1/p27(KIP1) levels in the cell.Perturbed integrin signaling may lead to the inhibition of SCF ligase activity, mislocalization and elevation of CKI-1/p27(KIP1).

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Baylor University, Waco, Texas, United States of America.

ABSTRACT
The cell-extracellular matrix (ECM) interaction plays an essential role in maintaining tissue shapes and regulates cell behaviors such as cell adhesion, differentiation and proliferation. The mechanism by which the ECM influences the cell cycle in vivo is poorly understood. Here we demonstrate that the β integrin PAT-3 regulates the localization and expression of CKI-1, a C. elegans homologue of the cyclin dependent kinase inhibitor p27(KIP1). In nematodes expressing wild type PAT-3, CKI-1::GFP localizes primarily to nucleoli in hypodermal cells, whereas in animals expressing mutant pat-3 with a defective splice junction, CKI-1::GFP appears clumped and disorganized in nucleoplasm. RNAi analysis links cell adhesion genes to the regulation of CKI-1. RNAi of unc-52/perlecan, ina-1/α integrin, pat-4/ILK, and unc-97/PINCH resulted in abnormal CKI-1::GFP localization. Additional RNAi experiments revealed that the SCF E3 ubiquitin-ligase complex genes, skpt-1/SKP2, cul-1/CUL1 and lin-23/F-box, are required for the proper localization and expression of CKI-1, suggesting that integrin signaling and SCF E3 ligase work together to regulate the cellular distribution of CKI-1. These data also suggest that integrin plays a major role in maintaining proper CKI-1/p27(KIP1) levels in the cell. Perturbed integrin signaling may lead to the inhibition of SCF ligase activity, mislocalization and elevation of CKI-1/p27(KIP1). These results suggest that adhesion signaling is crucial for cell cycle regulation in vivo.

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RNAi analysis showed that some focal adhesion and E3 ligase genes are required for CKI-1::GFP localization.Panels depict the results of RNAi of focal adhesion genes, dense body or M-line components, on the localization of CKI-1::GFP. Panels A: the negative control L4440 plasmid, B: pat-3 RNAi, C: ina-1 RNAi, D: pat-4 RNAi, E: unc-97 RNAi, F: pat-6 RNAi, G: unc-52 RNAi, H: let-2, I: skpt-1 RNAi, J: lin-23 RNAi and K: cul-1 RNAi.
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pone-0042425-g005: RNAi analysis showed that some focal adhesion and E3 ligase genes are required for CKI-1::GFP localization.Panels depict the results of RNAi of focal adhesion genes, dense body or M-line components, on the localization of CKI-1::GFP. Panels A: the negative control L4440 plasmid, B: pat-3 RNAi, C: ina-1 RNAi, D: pat-4 RNAi, E: unc-97 RNAi, F: pat-6 RNAi, G: unc-52 RNAi, H: let-2, I: skpt-1 RNAi, J: lin-23 RNAi and K: cul-1 RNAi.

Mentions: In pat-3(+) animals, pat-3(RNAi) resulted in CKI-1::GFP accumulation in the nucleoplasm similar to that seen in pat-3(sp) (Figures 5B and 2D). Next, integrin α subunits were depleted. Depletion of ina-1 in the pat-3(+) animals also resulted in abnormally clumped CKI-1::GFP (Figures 5C), suggesting that the CKI-1 localization is integrin dependent. Among the focal adhesion genes, pat-4/ILK [46], unc-97/PINCH [47] and pat-6/parvin [48] together form an IPP complex, which is implicated in the control of signaling pathways by the phosphorylation of downstream targets [49]. RNAi of pat-4/ILK or unc-97/PINCH in pat-3(+) resulted in the expected uncoordinated phenotypes (Figure S2) and CKI-1 mislocalization in hypodermal nuclei (Figure 5D and 5E). However, in pat-6/parvin RNAi animals, CKI-1 maintained its wild-type localization, possibly suggesting that the CKI-1 localization is independent of parvin (Figure 5F). Because pat-6 RNAi did not result in a strong uncoordinated phenotype (Figure S2), it is possible that the pat-6 RNAi is not as effective as the RNAi to pat-4 and unc-97. However, our data is consistent with the interpretation that ILK and PINCH are mediating integrin signals to control CKI-1 localization in the nucleus.


A novel mutation in β integrin reveals an integrin-mediated interaction between the extracellular matrix and cki-1/p27KIP1.

Kihira S, Yu EJ, Cunningham J, Cram EJ, Lee M - PLoS ONE (2012)

RNAi analysis showed that some focal adhesion and E3 ligase genes are required for CKI-1::GFP localization.Panels depict the results of RNAi of focal adhesion genes, dense body or M-line components, on the localization of CKI-1::GFP. Panels A: the negative control L4440 plasmid, B: pat-3 RNAi, C: ina-1 RNAi, D: pat-4 RNAi, E: unc-97 RNAi, F: pat-6 RNAi, G: unc-52 RNAi, H: let-2, I: skpt-1 RNAi, J: lin-23 RNAi and K: cul-1 RNAi.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3412830&req=5

pone-0042425-g005: RNAi analysis showed that some focal adhesion and E3 ligase genes are required for CKI-1::GFP localization.Panels depict the results of RNAi of focal adhesion genes, dense body or M-line components, on the localization of CKI-1::GFP. Panels A: the negative control L4440 plasmid, B: pat-3 RNAi, C: ina-1 RNAi, D: pat-4 RNAi, E: unc-97 RNAi, F: pat-6 RNAi, G: unc-52 RNAi, H: let-2, I: skpt-1 RNAi, J: lin-23 RNAi and K: cul-1 RNAi.
Mentions: In pat-3(+) animals, pat-3(RNAi) resulted in CKI-1::GFP accumulation in the nucleoplasm similar to that seen in pat-3(sp) (Figures 5B and 2D). Next, integrin α subunits were depleted. Depletion of ina-1 in the pat-3(+) animals also resulted in abnormally clumped CKI-1::GFP (Figures 5C), suggesting that the CKI-1 localization is integrin dependent. Among the focal adhesion genes, pat-4/ILK [46], unc-97/PINCH [47] and pat-6/parvin [48] together form an IPP complex, which is implicated in the control of signaling pathways by the phosphorylation of downstream targets [49]. RNAi of pat-4/ILK or unc-97/PINCH in pat-3(+) resulted in the expected uncoordinated phenotypes (Figure S2) and CKI-1 mislocalization in hypodermal nuclei (Figure 5D and 5E). However, in pat-6/parvin RNAi animals, CKI-1 maintained its wild-type localization, possibly suggesting that the CKI-1 localization is independent of parvin (Figure 5F). Because pat-6 RNAi did not result in a strong uncoordinated phenotype (Figure S2), it is possible that the pat-6 RNAi is not as effective as the RNAi to pat-4 and unc-97. However, our data is consistent with the interpretation that ILK and PINCH are mediating integrin signals to control CKI-1 localization in the nucleus.

Bottom Line: RNAi of unc-52/perlecan, ina-1/α integrin, pat-4/ILK, and unc-97/PINCH resulted in abnormal CKI-1::GFP localization.These data also suggest that integrin plays a major role in maintaining proper CKI-1/p27(KIP1) levels in the cell.Perturbed integrin signaling may lead to the inhibition of SCF ligase activity, mislocalization and elevation of CKI-1/p27(KIP1).

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Baylor University, Waco, Texas, United States of America.

ABSTRACT
The cell-extracellular matrix (ECM) interaction plays an essential role in maintaining tissue shapes and regulates cell behaviors such as cell adhesion, differentiation and proliferation. The mechanism by which the ECM influences the cell cycle in vivo is poorly understood. Here we demonstrate that the β integrin PAT-3 regulates the localization and expression of CKI-1, a C. elegans homologue of the cyclin dependent kinase inhibitor p27(KIP1). In nematodes expressing wild type PAT-3, CKI-1::GFP localizes primarily to nucleoli in hypodermal cells, whereas in animals expressing mutant pat-3 with a defective splice junction, CKI-1::GFP appears clumped and disorganized in nucleoplasm. RNAi analysis links cell adhesion genes to the regulation of CKI-1. RNAi of unc-52/perlecan, ina-1/α integrin, pat-4/ILK, and unc-97/PINCH resulted in abnormal CKI-1::GFP localization. Additional RNAi experiments revealed that the SCF E3 ubiquitin-ligase complex genes, skpt-1/SKP2, cul-1/CUL1 and lin-23/F-box, are required for the proper localization and expression of CKI-1, suggesting that integrin signaling and SCF E3 ligase work together to regulate the cellular distribution of CKI-1. These data also suggest that integrin plays a major role in maintaining proper CKI-1/p27(KIP1) levels in the cell. Perturbed integrin signaling may lead to the inhibition of SCF ligase activity, mislocalization and elevation of CKI-1/p27(KIP1). These results suggest that adhesion signaling is crucial for cell cycle regulation in vivo.

Show MeSH
Related in: MedlinePlus