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Identification of Plasmodesmal Localization Sequences in Proteins In Planta
Published on: August 15, 2017
Subcellular localization of MURA and MURB proteins encoded by the maize MuDR transposon
Akemi Ono1, Soo-Hwan Kim, Virginia Walbot
1Department of Biological Sciences, Stanford University, CA 94305-5020, USA
Abstract:
MuDR controls transposition of the Mu transposable element family in Zea mays L. It produces two major transcripts: mudrA and mudrB, mudrA encodes the MURA transposase, but no specific function has been ascribed to mudrB, which lacks strong homology to known genes. Using transient expression assays in onion epidermal cells, we defined three monopartite nuclear localization signals (NLSs) of MURA; each was functionally sufficient for nuclear targeting of MURA:GUS fusion proteins. Interestingly, one NLS (NLS-A3) is produced by the splicing of the third intron. In contrast, there were no clear NLS in MURB, and the major form of MURB aggregated in the cytoplasm. Self-interaction of MURA and of MURB was also shown in a yeast two-hybrid assay. To test whether interactions of MURA and MURB can occur at the level of protein translocation into the nucleus, a cytoplasmically localized MURB:GFP was co-expressed with MURA or with the GUS fusion proteins. Co-expression did not change the localization pattern of either MURA or MURB; MURA and MURB do not detectably interact in a yeast two-hybrid assay. These results suggest that MURA and MURB do not mutually affect their localization, at least in the forms examined here.
Insights
The MuDR element in maize produces MURA and MURB proteins. Researchers identified nuclear localization signals in MURA but not MURB, suggesting distinct cellular roles for these transposition proteins.
Area of Science:
- Plant molecular biology
- Genetics
- Transposable elements
Background:
- MuDR is a transposable element family in Zea mays (maize).
- MuDR produces two main transcripts: mudrA and mudrB.
- mudrA encodes the MURA transposase, essential for transposition.
Purpose of the Study:
- To investigate the cellular localization and potential interactions of MURA and MURB proteins.
- To identify nuclear localization signals (NLSs) in MURA and MURB.
- To determine if MURA and MURB influence each other's subcellular localization.
Main Methods:
- Transient expression assays in onion epidermal cells.
- Construction and testing of MURA:GUS and MURB:GFP fusion proteins.
- Yeast two-hybrid assays to assess protein-protein interactions.
Main Results:
- Three functional monopartite nuclear localization signals (NLSs) were identified in MURA, with one (NLS-A3) arising from intron splicing.
- No clear NLS were found in MURB, which predominantly localized to the cytoplasm.
- Co-expression of MURA and MURB did not alter their individual cellular localization patterns, and no interaction was detected in yeast two-hybrid assays.
Conclusions:
- MURA possesses functional NLSs enabling nuclear targeting, while MURB primarily resides in the cytoplasm.
- MURA and MURB do not appear to mutually influence each other's localization in the tested systems.
- The distinct localization patterns suggest separate functional roles for MURA and MURB in MuDR transposition.
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