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Characterization of the Multiple Domains of Pex30 Involved in Subcellular Localization of the Protein and Regulation of Peroxisome Number

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Abstract

Pex30 is a peroxisomal protein whose role in peroxisome biogenesis via the endoplasmic reticulum has been established. It is a 58 KDa multi-domain protein that facilitates contact site formation between various organelles. The present study aimed to investigate the role of various domains of the protein in its sub-cellular localization and regulation of peroxisome number. For this, we created six truncations of the protein (1-87, 1-250, 1-352, 88-523, 251-523 and 353-523) and tagged GFP at the C-terminus. Biochemical methods and fluorescence microscopy were used to characterize the effect of truncation on expression and localization of the protein. Quantitative analysis was performed to determine the effect of truncation on peroxisome number in these cells. Expression of the truncated variants in cells lacking PEX30 did not cause any effect on cell growth. Interestingly, variable expression and localization of the truncated variants in both peroxisome-inducing and non-inducing medium was observed. Truncated variants depicted different distribution patterns such as punctate, reticulate and cytosolic fluorescence. Interestingly, lack of the complete dysferlin domain or C-Dysf resulted in increased peroxisome number similar to as reported for cells lacking Pex30. No contribution of this domain in the reticulate distribution of the proteins was also observed. Our results show an interesting role for the various domains of Pex30 in localization and regulation of peroxisome number.

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Funding

This work was supported by the Department of Biotechnology (DBT), Government of India [BT/PR25097/NER/95/1013/2017] and Top-up of Start-Up grant from IIT Guwahati.

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Authors and Affiliations

  1. Organelle Biology and Cellular Ageing Lab, Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, 781039, India

    Nayan Moni Deori, Terence Infant & Shirisha Nagotu

  2. Laboratory for Stem Cell Engineering and Regenerative Medicine, Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, 781039, India

    Rajkumar P. Thummer

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Contributions

N.M.D. and T.I. performed the experiments. N.M.D. wrote the first draft of the manuscript and prepared the figures. R.P.T. analyzed the data and edited the manuscript. S.N. conceived the idea, analyzed the data, edited the manuscript and procured funding. All authors read and approved the final manuscript.

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Correspondence to Shirisha Nagotu.

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Deori, N.M., Infant, T., Thummer, R.P. et al. Characterization of the Multiple Domains of Pex30 Involved in Subcellular Localization of the Protein and Regulation of Peroxisome Number. Cell Biochem Biophys 81, 39–47 (2023). https://doi.org/10.1007/s12013-022-01122-z

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