Pon Lab Research Interests
Role of the actin cytoskeleton in control
of mitochondrial motility and inheritance in budding yeast:
Early studies of organelle inheritance focused almost exclusively
on the nucleus. These studies revealed that transfer of
the nucleus from mother to daughter cell is cytoskeleton-dependent
and required for cell division. It is now clear that similar
rules apply to the inheritance of other organelles. Our
studies focus on mitochondria, essential organelles that
must be produced from pre-existing mitochondria. We
identified a series of cell cycle-linked motility events
that lead to transfer of mitochondria from mother to daughter
cell, retention of newly inherited mitochondria in the bud,
and retention of remaining mitochondria in the mother cell.
These motility events are dependent upon the cytoskeleton,
the cell polarity machinery, and cell cycle regulatory pathways.
Thus, mitochondrial inheritance, like nuclear inheritance,
is an integral component of the cell division cycle.
We find that mitochondrial movement during yeast cell division
occurs by a mechanism that is similar, but not identical,
to that used by bacterial pathogens including Listeria monocytogenes
for movement through the cytoplasm of infected cells. Shigella
flexneri causes bacillary dysentery, a common disease with
worldwide distribution. Listeria monocytogenes is a food
borne pathogen responsible for meningitis, meningo-encephalitis,
and premature pregnancy termination with a mortality rate
of about 30%. Propulsion of these pathogens through the
cytoplasm of infected cells occurs by Arp2/3 complex-mediated
actin nucleation at a specific point along the bacterial
surface. This actin nucleation stimulates a burst of actin
polymerization, generating "actin comet tails"
that propel bacteria from one infected host cell to the
next. Mitochondrial movement, like that of the bacterial
pathogens, is dependent upon the Arp2/3 complex. However,
in contrast to bacterial patho-gens whose movement is randomly
directed, mitochondria use actin cables (bundles of actin
filaments that run along the mother-bud axis) as tracks
for movement from the mother cell to the bud.
According to our current working hypothesis the "mitochore"
is functionally homologous to the kinetochore and mediates
cyclic binding of mitochondrial membranes and mtDNA to actin
cables for movement in the presence of an applied force.
The force for movement is produced by: 1) Arp2/3 complex-stimulated
actin polymerization and assembly at the interface between
mitochondria and actin cables, and 2) cross-linking of mitochondria-associated
actin polymers in parallel to F-actin within actin cables.
The net effect of these processes is movement of mitochondria
towards the barbed or fast-growing end of actin cables,
and linear, polarized movement of mitochondria from mother
to daughter cells.
Our efforts to understand this process include molecular
dissection of the proteins that 1) mediate association of
mitochondria with the actin cytoskeleton and with the Arp2/3
complex, and 2) regulate mitochondrial movement and inheritance
during cell division. Given the fundamental role of mitochondria
in aerobic energy mobilization and biosynthesis of molecules
including amino acids, fatty acids, pyrimidines, heme and
steroid hormones, it is surprising the so little is known
regarding the mechanisms responsible for transmission of
mitochondria from mother to daughter cell during cell division.
Our studies on mitochondrial movement and inheritance are
designed to fill that gap.
Actin dynamics during establishment of
cell polarity in budding yeast:
Actin cables are bundles of filamentous actin, which align
along the mother-bud axis and resemble actin bundles in
mammalian cells (e.g. the bristle and follicular nurse cell
struts of Drosophila, the brush border of intestinal epithelial
cells, and the stereocilium of hair cells in the vertebrate
ear). Studies with tropomyosin mutants, which show temperature-dependent
loss of actin cables, implicate actin cables as tracks for
movement of components from mother cell to bud including
secretory vesicles, mRNA, lysomes, mitochondria, and elements
required for spindle alignment. Despite the fundamental
role of these structures in polarized growth and yeast cell
division, the events associated with actin cable polarization
and maintenance are not well understood. We
visualized actin cables in living yeast using fluorescent
tags and time lapse fluorescence micrscopy. We find that
actin cables are dynamic structures that undergo elongation
and cortical movement, and achieve their polarization by
assembly and extension along the mother-bud axis. Moreover,
we localized sites of actin cable assembly within the incipient
bud, bud and bud neck. Future studies will focus on analysis
of the role of known actin binding proteins and polarity
factors in actin cable assembly, movement and disassembly.
These studies will extend our understanding of other fundamental
actin-dependent motility events including 1) vesicle movement
during secretion and neuronal transmission, 2) development
and maintenance of apical modifications in lymphocytes,
intestinal brush border cells, migrating neurons and other
cells, 3) asymmetric distribution of organelles and mRNA
during development, and 4) polarized cell growth during
host invasion by pathogenic fungi (e.g. Candida albicans,
Aspergillus nidulans) responsible for human infections including
thrush, vaginal candidiasis, skin infections, diaper rash,
and esophagitis, as well as intestinal, urinary, central
nervous system, cardiac and disseminated infections.