Molecular Biology Commons^™

Rapid Screening For Temperature-Sensitive Alleles In Plants, L Vidali, Rc Augustine, Sn Fay, P Franco, Ka Pattavina, M Bezanilla

Magdalena Bezanilla

We developed a simple and fast method to identify temperature-sensitive alleles of essential plant genes. We used primary and tertiary structure information to identify residues in the core of the protein of interest. These residues were mutated and tested for temperature sensitivity, taking advantage of the exceptionally rapid 1-week complementation assay in the moss Physcomitrella patens. As test molecules, we selected the actin-binding proteins profilin and actin-depolymerizing factor, because they are essential and their loss-of-function phenotype can be fully rescued. Screening a small number of candidate mutants, we successfully identified temperature-sensitive alleles of both profilin and actin-depolymerizing factor. Plants harboring …

Lifeact-Megfp Reveals A Dynamic Apical F-Actin Network In Tip Growing Plant Cells, L Vidali, Cm Rounds, Pk Hepler, M Bezanilla

Magdalena Bezanilla

Background

Actin is essential for tip growth in plants. However, imaging actin in live plant cells has heretofore presented challenges. In previous studies, fluorescent probes derived from actin-binding proteins often alter growth, cause actin bundling and fail to resolve actin microfilaments.

Methodology/Principal Findings

In this report we use Lifeact-mEGFP, an actin probe that does not affect the dynamics of actin, to visualize actin in the moss Physcomitrella patens and pollen tubes from Lilium formosanum and Nicotiana tobaccum. Lifeact-mEGFP robustly labels actin microfilaments, particularly in the apex, in both moss protonemata and pollen tubes. Lifeact-mEGFP also labels filamentous actin structures in …