Functional diversity of Medicago truncatula RNA polymerase II CTD phosphatase isoforms produced in the Arabidopsis thaliana superexpression platform

Akihito Fukudome; Yasuhiro Ishiga; Yukihiro Nagashima; Katherine H Davidson; Hsiu-An Chou; Kirankumar S Mysore; Hisashi Koiwa

doi:10.1016/j.plantsci.2022.111309

Functional diversity of Medicago truncatula RNA polymerase II CTD phosphatase isoforms produced in the Arabidopsis thaliana superexpression platform

Plant Sci. 2022 Aug:321:111309. doi: 10.1016/j.plantsci.2022.111309. Epub 2022 May 10.

Authors

Akihito Fukudome¹, Yasuhiro Ishiga², Yukihiro Nagashima³, Katherine H Davidson³, Hsiu-An Chou¹, Kirankumar S Mysore⁴, Hisashi Koiwa⁵

Affiliations

¹ Molecular and Environmental Plant Sciences, Texas A&M University, College Station, TX 77843, USA.
² Noble Research Institute, LLC., Ardmore, OK 73401, USA; Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan.
³ Vegetable and Fruit Development Center, Department of Horticultural Sciences, Texas A&M University, College Station, TX 77843, USA.
⁴ Noble Research Institute, LLC., Ardmore, OK 73401, USA; Institute for Agricultural Biosciences, Oklahoma State University, Ardmore, Oklahoma 73401, USA; Department of Biochemistry and Molecular Biology, Oklahoma State University, Stillwater, Oklahoma 74044, USA.
⁵ Molecular and Environmental Plant Sciences, Texas A&M University, College Station, TX 77843, USA; Vegetable and Fruit Development Center, Department of Horticultural Sciences, Texas A&M University, College Station, TX 77843, USA. Electronic address: hisashi.koiwa@ag.tamu.edu.

PMID: 35696909
DOI: 10.1016/j.plantsci.2022.111309

Abstract

Medicago truncatula is a model system for legume plants, which has substantially expanded the genome relative to the prototypical model dicot plant, Arabidopsis thaliana. An essential transcriptional regulator, FCP1 (transcription factor IIF-interacting RNA polymerase II carboxyl-terminal phosphatase 1) ortholog, is encoded by a single essential gene CPL4 (CTD-phosphatase-like 4), whereas M. truncatula genome contains four genes homologous to FCP1/AtCPL4, and splicing variants of MtCPL4 are observed. Functional diversification of MtCPL4 family proteins was analyzed using recombinant proteins (MtCPL4a1, MtCPL4a2, and MtCPL4b) produced in Arabidopsis cell culture system developed for plant protein overexpression. In vitro CTD phosphatase assay using highly purified MtCPL4 preparations revealed a potent CTD phosphatase activity in MtCPL4b, but not two splicing variants of MtCPL4a. On the other hand, in planta binding assay to RNA polymerase II (pol II) revealed a greater pol II-binding activity of both MtCPL4a variants. Our results indicate functional diversification of MtCPL4 isoforms and suggest the presence of a large number of functionally specialized CTD-phosphatase-like proteins in plants.

Keywords: Arabidopsis thaliana; CTD phosphatase; Medicago truncatula; Plant recombinant protein; RNA polymerase II.

MeSH terms

Arabidopsis Proteins* / genetics
Arabidopsis* / genetics
Arabidopsis* / metabolism
Medicago truncatula* / genetics
Medicago truncatula* / metabolism
Phosphoprotein Phosphatases / genetics
Protein Isoforms / genetics
RNA Polymerase II / genetics
RNA Polymerase II / metabolism

Substances

Arabidopsis Proteins
Protein Isoforms
RNA Polymerase II
CPL4 protein, Arabidopsis
Phosphoprotein Phosphatases
carboxy-terminal domain phosphatase