In fungi, cell shape is determined by the presence of a rigid cell wall whi
ch separates the cell from the extracellular medium. This highly dynamic st
ructure is essential for the maintenance of cell integrity and is involved
in several phenomena such as flocculation, adherence and pathogenicity. The
composition of the fungal cell wall is well known, but issues such as the
assembly and remodeling of its components remain poorly understood. In an a
ttempt to study the de novo construction of the yeast cell wall, we have un
dertaken a large-scale proteomic approach to analyze the proteins secreted
by regenerating protoplasts. Upon incubation of protoplasts in regenerating
conditions, numerous proteins are secreted into the culture medium. These
presumably include proteins destined for the cell wall, comprising both str
uctural proteins as well as enzymes involved in cell wall biogenesis. This
work reports the establishment of a reference map of proteins secreted by r
egenerating protoplasts by means of two-dimensional polyacrylamide gel elec
trophoresis (2-D PAGE) and their identification by mass spectrometry. Thirt
y-two different proteins have been identified, including known cell wall pr
oteins, glycolytic enzymes, heat shock proteins, and proteins involved in s
everal other processes. Using this approach, novel proteins possibly involv
ed in cell wall construction have also been identified. This reference map
will allow comparative analyses to be carried out on a selected collection
of mutants affected in the cell wall.