Polar Growth in Streptomyces venezuelae

Abstract: Cell polarity is an ubiquitous and important phenomenon relevant for a plethora of cellular functions. One type of cell polarity is polar growth, where an organism only synthesizes cell envelope at designated locations in the cell. This requires highly organized localization of cellular activities. Understanding the ways in which polarity and polar growth are organized is critical for understanding how cell shape is established and maintained. Many industrial or clinically important organisms exhibit polar growth, making studies thereof widely relevant.The overall aim of this thesis was to investigate polar growth using, the Gram-positive actinomycete Streptomyces venezuelae as a model. In the filamentous S. venezuelae, polar growth is organized by the polarisome. The polarisome is a protein complex responsible for the recruitment and maintenance of the cell wall synthesis machinery at hyphal tips of the vegetative mycelium.One part of the work presented here focuses on FilP, a protein known to affect polar growth. Our investigations of FilP clarify its localization in the cell, its function, and establish that it is a very dynamic protein. We also show that the localization of a fluorescent fusion protein of FilP is dependent on active hyphal extension. We establish that FilP affects the polarisome protein DivIVA, a protein throught to be the main organizer of polar growth in Streptomyces, likely leading to the observed tortuous morphology of the filP deletion mutant strain. We also show that FilP localization is not strictly dependent on the polarisome protein Scy.The work presented in this thesis, establishes details of the polar growth synthesis machinery. The work also establishes that the cell wall synthesis activities, represented by labeling with both fluorescently labeled vancomycin and fluorescent D-amino acids, localize to the apex of the hyphae in Streptomyces. We also report that FilP is likely required for normal localization of the cell wall synthesis machinery.In this thesis, a new phenomenon of vesicle extrusion from vegetative hyphal tips of Streptomyces is reported. We establish that the vesicle extrusion is coupled to reprogramming of polar growth and is likely related to the polarisome and polar growth. We identify a genetic determinant, scy, that protects from vesicle extrusion. This may be, to the best of our knowledge, the first demonstrated vesicle extrusion not leading to cell death in a Gram-positive organism.This thesis aims to present my work, discuss my findings and put my work into perspective.