Abstract

Functional studies of yeast mutants reveal mechanisms for TRAPP-complex integrity

Stephanie Brunet, PhD
Concordia University, 2015

Proteins and lipids are packaged into membrane bound vesicles and transported
throughout the cell in a highly regulated process called membrane trafficking. Several
categories of proteins including tethering factors, cooperate to ensure fidelity in this
process that function as a physical a link between a vesicle and the correct target
membrane. The transport protein particle (TRAPP) complex is a multisubunit tethering
complex that has been well characterized in yeast and is highly conserved in mammals. In
yeast, three isoforms of the complex (TRAPPI, TRAPPII and TRAPPIII) have been described
each acting in different membrane trafficking pathways. My research has focused on
studying yeast TRAPP mutants to understand how specific subunits contribute to TRAPPcomplex
integrity and how each complex regulates distinct membrane trafficking
pathways.
Several mutations in the human TRAPPC2 gene including a missense mutation of a
highly‐conserved aspartic acid residue (TRAPPC2D47Y), are linked to the skeletal disorder
spondyloepiphyseal dysplasia tarda (SEDT), Chapter 2 discusses how the analogous
mutation in the yeast homologue, TRS20, destabilizes the TRAPPIII complex and disrupts
TRAPPIII‐regulated membrane trafficking pathways, providing insight into the etiology of
SEDT.
Chapter 3 of this thesis describes a mutational study that I conducted on the
essential yeast TRAPP subunit, Trs23p. When a non‐essential Saccharomycotina‐specific
(SMS) domain within this protein is deleted, TRAPPI, the smallest of the complexes, is
destabilized however processes regulated by TRAPPI are not compromised. TRAPPII and
TRAPPIII contain all the subunits of the smaller TRAPPI complex and I propose that
TRAPPI is a fragment or assembly intermediate of the larger complexes.
Finally, in Chapter 4 I present evidence for a direct interaction between the TRAPPII
complex and Gyp6p, a GTPase accelerating protein (GAP) for the small Rab GTPase Ypt6p.
In mutants that destabilize the TRAPPII complex this interaction is disrupted and Ypt6p
localization to the late Golgi compartment is increased relative to wild type. Together these
results indicate that the interaction between TRAPPII and Gyp6p is important for
regulating the dynamic distribution of Ypt6p within the cell.