Evaluating the activity of the filamentous growth mitogen-activated protein kinase pathway in yeast

Cold Spring Harbor Protocols
Paul J Cullen

Abstract

Mitogen-activated protein kinase (MAPK) pathways are evolutionarily conserved signaling pathways that regulate diverse processes in eukaryotes. One such pathway regulates filamentous growth, a nutrient limitation response in budding yeast and other fungal species. This protocol describes three assays used to measure the activity of the filamentous growth pathway. First, western blotting for phosphorylated (activated) MAPKs (P∼MAPKs; Slt2p, Kss1p, Fus3p, and Hog1p) provides a measure of MAPK activity in yeast and other fungal species. Second, the PGU1 gene is a transcriptional target of the filamentous growth pathway. Cells that undergo filamentous growth secrete Pgu1p, an endopolygalacturonase that degrades the plant-specific polysaccharide pectin. We describe an assay that measures secreted pectinase activity, which reflects an active filamentous growth pathway. Finally, in yeast, two mucin-like glycoproteins, Msb2 and Flo11, regulate filamentous growth. Secretion of the processed and shed glycodomain of Msb2 is an indicator of MAPK activity. Flo11, the major adhesion molecule that controls filamentous growth and biofilm/mat formation, is also shed from cells. Detecting shed mucins with epitope-tagged versions of the proteins ...Continue Reading

References

Oct 27, 1999·Proceedings of the National Academy of Sciences of the United States of America·H D MadhaniG R Fink
Feb 12, 2008·Current Biology : CB·Michael J Lee, Henrik G Dohlman
Mar 25, 2010·PLoS Genetics·Colin A ChavelPaul J Cullen
Jul 14, 2010·Current Biology : CB·Sheelarani KarunanithiPaul J Cullen
Mar 29, 2011·The Plant Cell·Elena Pérez-Nadales, Antonio Di Pietro
Feb 10, 2012·PLoS Pathogens·Eva Szafranski-SchneiderJoachim F Ernst
Aug 30, 2012·Nature Methods·Caroline A SchneiderKevin W Eliceiri

❮ Previous
Next ❯

Citations

Mar 4, 2015·Cold Spring Harbor Protocols·Paul J Cullen

❮ Previous
Next ❯

Related Concepts

Related Feeds

Biofilm & Infectious Disease

Biofilm formation is a key virulence factor for a wide range of microorganisms that cause chronic infections.Here is the latest research on biofilm and infectious diseases.

Adhesion Molecules in Health and Disease

Cell adhesion molecules are a subset of cell adhesion proteins located on the cell surface involved in binding with other cells or with the extracellular matrix in the process called cell adhesion. In essence, cell adhesion molecules help cells stick to each other and to their surroundings. Cell adhesion is a crucial component in maintaining tissue structure and function. Discover the latest research on adhesion molecule and their role in health and disease here.

Biofilms

Biofilms are adherent bacterial communities embedded in a polymer matrix and can cause persistent human infections that are highly resistant to antibiotics. Discover the latest research on Biofilms here.