Identification of a truncated form of methionine sulfoxide reductase a expressed in mouse embryonic stem cells

Pingping Jia,Keith A Webster,Xupei Huang,Chi Zhang,Andrei A Kochegarov,Larry F Lemanski,Sharon L Lemanski,Yuanyuan Jia

doi:10.1186/1423-0127-18-46

Abstract

BackgroundMethionine Sulfoxide Reductase A (MsrA), an enzyme in the Msr gene family, is important in the cellular anti-oxidative stress defense mechanism. It acts by reducing the oxidized methionine sulfoxide in proteins back to sulfide and by reducing the cellular level of reactive oxygen species. MsrA, the only enzyme in the Msr gene family that can reduce the S-form epimers of methionine sulfoxide, has been located in different cellular compartments including mitochondria, cytosol and nuclei of various cell lines.MethodsIn the present study, we have isolated a truncated form of the MsrA transcript from cultured mouse embryonic stem cells and performed eGFP fusion protein expression, confocal microscopy and real time RT-PCR studies.ResultsResults show a different expression response of this truncated transcript to oxygen deprivation and reoxygenation treatments in stem cells, compared to the longer full length form. In addition, a different subcellular localization pattern was noted with most of the eGFP fusion protein detected in the cytosol.ConclusionOne possibility for the existence of a truncated form of the MsrA transcripts could be that with a smaller protein size, yet retaining a GCWFG action site, this protein might have easier access to oxidize methionine residues on proteins than the longer form of the MsrA protein, thus having an evolutionary selection advantage. This research opens the door for further study on the role and function of the truncated MsrA embryonic mouse stem cells.

Highlights

Methionine Sulfoxide Reductase A (MsrA), an enzyme in the Msr gene family, is important in the cellular anti-oxidative stress defense mechanism
There are protein bands with molecular weights of ~46~48 kD indicating homodimers from MsrA long form proteins (Figure 1A, arrow head), even with protein samples thoroughly treated by reducing agents before SDS-PAGE, consistent with findings from other laboratories [10]
There is a band with a molecular weight of 39 kD, possibly hetero-dimers formed by a long form protein of MsrA (MW: 23 kD) and a smaller form of MsrA protein (MW 16 kD) (Figure 1A, *)

Summary

Introduction

Methionine Sulfoxide Reductase A (MsrA), an enzyme in the Msr gene family, is important in the cellular anti-oxidative stress defense mechanism. MsrA, the only enzyme in the Msr gene family that can reduce the S-form epimers of methionine sulfoxide, has been located in different cellular compartments including mitochondria, cytosol and nuclei of various cell lines. One of the cellular defense mechanisms to reduce the oxidized residues in proteins, and restore their functions as well as reduce oxidative stress, relies on the methionine sulfoxide reductase (msr) family genes, of which three MsrB genes (MsrB1, B2 and B3), have been identified in mammals [1,2,3,4,5,6,7]. Human MsrB3 gives rise to two forms generated by alternative first exon splicing, which are targeted to the endoplasmic reticulum (ER) and mitochondria [4] of the cell

Methods

Results

Conclusion