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. 2019 Aug;20(2):1790-1796.
doi: 10.3892/mmr.2019.10419. Epub 2019 Jun 24.

Identification of COMMD1 as a Novel Lamin A Binding Partner

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Free PMC article

Identification of COMMD1 as a Novel Lamin A Binding Partner

Zhiwen Jiang et al. Mol Med Rep. .
Free PMC article

Abstract

Lamin A, which is encoded by the LMNA gene, regulates gene expression and genome stability through interactions with a variety of proteins. Mutations in LMNA lead to a diverse set of inherited human diseases, collectively referred to as laminopathies. To gain insight into the protein interactions of lamin A, a yeast two‑hybrid screen was conducted using the carboxy‑terminus of lamin A. The screen identified copper metabolism MURR1 domain‑containing 1 (COMMD1) as a novel lamin A binding partner. Colocalization experiments using fluorescent confocal microscopy revealed that COMMD1 colocalized with lamin A in 293 cells. Furthermore, the COMMD1‑lamin A protein interaction was also demonstrated in co‑immunoprecipitation experiments. Collectively, the present study demonstrated a physical interaction between COMMD1 and lamin A, which may aid to elucidate the mechanisms of lamin A in the aging process.

Figures

Figure 1.
Figure 1.
Analysis of the COMMD1-lamin A interaction using a yeast two-hybrid assay. (A) Positive clones interacting with the C-terminus of lamin A on low-stringency synthetic defined/histidine/leucine/tryptophan plates containing X-α-gal (blue). (B) Schematic diagram of the interaction between the COMMD1 sequence and the C-terminus of lamin A. (C) Schematic diagram of the second round selection using a high stringency protocol. (D) Diploid AH109 yeast were re-streaked onto synthetic defined/adenine/histidine/leucine/tryptophan plates containing X-α-gal to test for an interaction. AH109 yeast transformed with the C-terminus of lamin A in the pGBKT7 vector. Empty pGBKT7-LA-C was used as a negative control. AH109 yeast transformed with pCL1 was used as positive control. A blue signal indicates the activation of the reporter genes. COMMD1, copper metabolism MURR1 domain-containing 1; C-terminus, carboxy-terminus; X-α-gal, 5-bromo-4-chloro-3-indolyl-α-D-galactopyranoside.
Figure 2.
Figure 2.
Co-localization of COMMD1 and lamin A in 293 cells. Co-transfection of 293 cells with (A) GFP-COMMD1 and pDsRed1-N1 (RFP) or (B) GFP-COMMD1 and RFP-lamin A. (C) Higher magnification view of cells co-transfected with GFP-COMMD1 and pDsRed1-N1-lamin A (RFP-lamin A). Cells were examined 24 h following transfection using confocal scanning laser microscopy. COMMD1, copper metabolism MURR1 domain-containing 1; GFP, green fluorescent protein; RFP, red fluorescent protein; RFP-lamin A, pDsRed1-N1-lamin A.
Figure 3.
Figure 3.
COMMD1 interacts with lamin A. (A) IP of 293 cell extracts with anti-lamin A/C or anti-COMMD1 antibodies, followed by western blotting with anti-COMMD1 and anti-lamin A/C antibodies. (B) Following co-transfection of 293 cells with pCMV-HA or HA-COMMD1 and Flag-lamin A, IP was conducted with an anti-HA antibody, followed by western blotting using an anti-Flag antibody. COMMD1, copper metabolism MURR1 domain-containing 1; HA, hemagglutinin; Ab, antibody; IgG, immunoglobulin G; WCE, whole cell extract; IP, immunoprecipitation; WB, western blotting.

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