Molecular Genetics of Intracellular 
Protein Sorting 
Scott D. Emr, Ph.D. — Associate Investigator 
Dr. Emr is also Professor of Cellular and Molecular Medicine at the University of California, San Diego, 
School of Medicine. He did his graduate work in microbiology and molecular genetics with Thomas 
Silhavy and Jonathan Beckwith at Harvard Medical School, where he identified and characterized the first 
secretion- defective signal sequence mutants as well as the first component of the bacterial protein export 
apparatus. Dr. Emr did postdoctoral work on protein secretion in yeast with Randy Schekman at the 
University of California, Berkeley. He was a faculty member at the California Institute of Technology 
before moving to UCSD. Dr. Emr counts among his honors a Searle Scholars Award and an NSF 
Presidential Young Investigator Award. 
AN essential feature of all eukaryotic cells is 
their highly compartmentalized organiza- 
tion. Different, often competing biochemical 
processes are segregated into distinct compart- 
ments. The identity, stability, and function of 
each of these compartments, or organelles, is 
conferred in large part by the unique set of pro- 
teins that reside within it. These proteins must be 
routed from their common site of synthesis in the 
cytoplasm to their unique site of function in the 
appropriate intracellular organelle. Toward a de- 
tailed understanding of the molecular mecha- 
nisms that direct the delivery of one class of these 
proteins, we have focused our attention on the 
transport and sorting of proteins through the 
Golgi complex to the lysosome. 
Lysosomal Hydrolase Sorting 
Our laboratory is using a simple unicellular eu- 
karyote, the yeast Saccharomyces cerevisiae, as a 
model genetic system to study protein trafficking 
through the secretory pathway to the lysosome- 
like vacuole. The fundamental similarities be- 
tween yeast and other eukaryotic cells in their 
pathways and mechanisms for protein delivery 
have clearly established yeast as an important 
model system for the study of these problems. 
The importance of the lysosomal protein sort- 
ing pathway in humans is revealed when the ef- 
fects of mislocalizing such enzymes are exam- 
ined. A number of lysosomal hydrolases are 
secreted from naturally occurring tumor cells 
(e.g., cathepsin D by breast cancer cells). It has 
been proposed that the mislocalization of lyso- 
somal enzymes enhances cell growth and in- 
creases the metastatic potential of tumor cells by 
contributing to the hydrolysis of extracellular 
matrix components of target tissues. The impor- 
tance of this sorting pathway is further exempli- 
fied by the serious inherited disorders (e.g., I cell 
disease) that result in mislocalization of lyso- 
somal hydrolases. 
Proteins destined for the vacuole/lysosome in 
yeast and mammalian cells transit through early 
stages of the secretory system. The transport and 
processing characteristics of several vacuolar hy- 
drolases in yeast, including the soluble protease 
carboxypeptidase Y (CPY), have been well char- 
acterized. CPY is translocated into the endoplas- 
mic reticulum (ER), where it is modified with 
four core oligosaccharides to generate the ER 
precursor form of CPY (pi CPY). Subsequent 
transport events are mediated by vesicular car- 
riers. Delivery from the ER to the Golgi complex 
and passage through the Golgi are accompanied 
by elongation of the core oligosaccharides on 
CPY, resulting in formation of the Golgi- 
modified form of CPY (p2CPY). Sorting of 
p2CPY from proteins destined for secretion ap- 
pears to take place in the late Golgi. Upon arrival 
in the vacuole, an amino-terminal propeptide 
segment on p2CPY is proteolytically removed, 
generating the active mature form of the protease 
(mCPY). 
Sorting-Defective Mutants 
We have focused our efforts on identifying and 
characterizing the cellular machinery that directs 
the sorting and transport of vacuolar hydrolases. 
A gene fusion-based selection scheme we de- 
signed enabled us to isolate more than 600 yeast 
mutants that exhibit severe defects in vacuolar 
protein sorting. Each of the mutants missorts and 
secretes CPY and other vacuolar enzymes. The 
recessive mutations define more than 33 comple- 
mentation groups referred to as vps (vacuolar 
protein sorting defective) . Extensive genetic, bio- 
chemical, and morphological characterization of 
the vps mutants has allowed us to organize them 
into phenotypically related groups. Each group 
appears to function at a common stage of the pro- 
tein sorting pathway. 
The apparent genetic complexity of the vps 
mutant collection presumably reflects the bio- 
chemical complexity of the protein sorting reac- 
tion. Specific cellular components must recog- 
nize vacuolar proteins, segregate them from 
other secretory proteins, and package them into 
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