Analysis of Organelle Targeting by DIL Domains of the Arabidopsis Myosin XI Family

Amirali Sattarzadeh,Elmon Schmelzer,Maureen R Hanson

doi:10.3389/fpls.2011.00072

Abstract

The Arabidopsis thaliana genome encodes 13 myosin XI motor proteins. Previous insertional mutant analysis has implicated substantial redundancy of function of plant myosin XIs in transport of intracellular organelles. Considerable information is available about the interaction of cargo with the myosin XI-homologous yeast myosin V protein myo2p. We identified a region in each of 12 myosin XI sequences that correspond to the yeast myo2p secretory-vesicle binding domain (the “DIL” domain). Structural modeling of the myosin DIL domain region of plant myosin XIs revealed significant similarity to the yeast myo2p and myo4p DIL domains. Transient expression of YFP fusions with the Arabidopsis myosin XI DIL domain resulted in fluorescent labeling of a variety of organelles, including the endoplasmic reticulum, peroxisomes, Golgi, and nuclear envelope. With the exception of the YFP::MYA1 DIL fusion, expression of the DIL–YFP fusions resulted in loss of motility of labeled organelles, consistent with a dominant-negative effect. Certain fusions resulted in localization to the cytoplasm, plasma membrane, or to unidentified vesicles. The same YFP-domain fusion sometimes labeled more than one organelle. Expression of a YFP fusion to a yeast myo2p DIL domain resulted in labeling of plant peroxisomes. Fusions with some of the myosin XI domains resulted in labeling of known cargoes of the particular myosin XI; however, certain myosin XI YFP fusions labeled organelles that had not previously been found to be detectably affected by mutations nor by expression of dominant-negative constructs.

Highlights

Arabidopsis thaliana and other vascular plants encode genes for myosin motor proteins involved in vesicle transport (Reddy, 2001)
MYOSIN XI DOMAINS HOMOLOGOUS TO THE YEAST SECRETORY-VESICLE-SPECIFIC DOMAINS The class V myosin Myo2p in S. cerevisiae harbors within its C-terminal tail region two distinct regions required for cargobinding (Pashkova et al, 2006). These regions were identified by mutagenesis of the tail domain of yeast Myo2p followed by measurement of the ability of mutated Myo2p to carry the different organelles to the bud (Catlett et al, 2000; Pashkova et al, 2005)
Arabidopsis myosin XI proteins contain a domain that is homologous to the yeast myosin V domain that was shown to be important for secretory-vesicle movement (Figure 1)

Summary

Introduction

Arabidopsis thaliana and other vascular plants encode genes for myosin motor proteins involved in vesicle transport (Reddy, 2001). The plant myosin XI family is evolutionarily related to the wellstudied myosin V family in yeast and mammals (Kinkema and Schiefelbein, 1994). Both myosin V and XI proteins carry a number of different domains that are required for motor activity, binding of light chains, dimerization, or attachment of cargo (Li and Nebenfuhr, 2007). Single Arabidopsis myosin gene insertional mutants have been reported to exhibit little whole-plant phenotype, implying considerable redundancy of function (Ojangu et al, 2007; Peremyslov et al, 2008). Movement of certain organelles has been observed to be impaired in mutants or in cells expressing defective myosins or in which myosin gene silencing has occurred (Ojangu et al, 2007; Peremyslov et al, 2008, 2010; Avisar et al, 2009)

Objectives

Methods

Results

Conclusion