LT16 Regulation of Cell Growth

Regulation of Cell Growth

 Cell growth is an increase in cell mass and size
 Proliferation is an increase in cell number

 Size of cell depends on intrinsic and extrinsic factors – cell size can vary
dramatically with cell type – some neurons or glia cells are up to 1000 times larger
(Hafen + Stocker, 2003)
 Cell size is influenced by number of genome sets (ploidy)
 Amongst extrinsic factors – availability of nutrients and temperature
Eg. starvation prolongs cell doubling time in yeast and in Drosophila cells
 In yeast – evidence for a cell size checkpoint (Nurse 1975)
Budding yeast - protein Cln3p (G1 cyclin) acts as a sizer – Cells only initiate critical
cell cycle step from G1 phase to S phase when Cln3p has reached a certain
threshold
Accumulation of Cln3p is dependent on efficient translation of Cln3 mRNA, which is
inefficiently translated until sufficient numbers of ribosomes have been generated
Yeast size mutants exhibited defects in ribosome biogenesis (Jorgensen et al.,
2002)


Mitogens stimulate cell division

 Mitogens are secreted signal proteins that bind to cell-
surface receptors
 When activated by mitogen binding, these receptors
initiate various intracellular signalling pathways that
stimulate cell division – act mainly by releasing molecular
brakes that block transition from G1 phase of the cell
cycle into S phase
 Retinoblastoma protein (Rb) – abundant in nucleus of
vertebrate cells – binds to particular transcript regulators
preventing stimulation of the transcription of genes
required for cell proliferation
 Platelet-derived growth factor (PGDF) – PGDF binds to
receptor tyrosine kinases in surviving cells at wound site.
Which stimulate them to proliferate and heal wound
 Hepatocyte growth factor – stimulate surviving liver cells
to proliferate




Nutrients + ATP are required for biosynthesis

,Transport of nutrients done by:

- Receptor mediated endocytosis
- Phagocytosis
- Proteasomal degradation
- Autophagy
- Amino acid transporters
- Glucose transporters
- Ion channels
- Nutrient receptors = LDL-R, TfR

Regulation of cell growth by:

Growth Factors + Hormones

 Insulin, IGF1, PDGF-R, NGF
 Similar to mitogens, most extracellular growth factors bind to cell-
surface receptors, activating various intracellular signalling
pathways, lea to accumulation of proteins and other macromolecules
 Do so by increasing rate of synthesis of molecules and decreasing
rate of degradation
 Some signal proteins (eg. PDGF) act as both growth factors and
mitogens – stimulating both cell growth and progression through cell
cycle
 Insulin and insulin-like growth factor receptors (IR or IGFR) key regulators of
growth signalling
- IR is a heterotetramer
- Insulin binding induces conformation changes and stimulation of receptor
tyrosine kinase activity
- IR autophosphorylates and phosphorylates downstream second messengers (eg,
Insulin Receptor Substrate (IRS))
 Signalling via Receptor Tyrosine Kinases

Cytokines

 Interleukins
 Signalling via tyrosine kinase-associated receptors

Steroids

 Signalling via nuclear receptors

mTOR = mammalian target of rapamycin signalling pathway

, Signalling pathway integrates both intracellular and
extracellular signals serves as a central regulator of cell
metabolism, growth, proliferation and survival
 mTOR protein is a Ser/Thr protein kinase (289kDa)-
nucleates at least 2 distinct multi-protein complexes
(Laplante and Sabatini, 2009)
 mTOR sense availability of amino acids, metabolic fuel and
energy
 Nutrients and energy stores are essential for protein
synthesis, cell growth, proliferation and survival

mTOR forms 2 regulatory complexes in cells

mTORC1

 Composed of mTOR (catalytic subunit), regulator-
associated protein of mTOR (Raptor), mammalian
lethal with Sec13 protein 8 (GβL)
 Positively regulates cell growth and proliferation
by promoting many anabolic processes, including
biosynthesis of proteins, lipids and organelles, and limit catabolic processes such as
autophagy
 Much of knowledge about mTORC1 function comes from use of bacterial macrolide
rapamycin
- Rapamycin interacts with the FKBP12-rapamycin binding domain (FRB) of mTOR,
thus inhibiting mTORC1 functions (Guertin and Sabatini, 2007)
- FKB12-rapamycin cannot physically interact with or acutely inhibit mTORC2
(Jacinto et al., 2004)
- mTORC1 is rapamycin-sensitive and mTORC2 is rapamycin-insensitive
- Chronic rapamycin treatment in some cases can inhibit mTORC2 by blocking its
assembly (Choo et al., 2008)
 mTORC1 integrates 4 major signals – growth factors, energy status, oxygen and
amino acids to regulate cell growth
- Energy status – signalled to mTORC1 through AMP-activated protein kinase
(AMPK) – master sensory of intracellular energy status (Hardie, 2007), responds
to low ATP:ADP ratio
- Oxygen levels – under conditions of mild hypoxia – reduction in ATP levels
activates AMPK, which inhibits mTORC1 signalling
- Amino acids – represent strong signal that positively regulates mTORC1

, Rag proteins (family of 4 related small
GTPases) interact with mTORC1 in an
amino acid-sensitive manner
Rag proteins bind to Raptor and promote
the relocalisation of mTORC1 from
discrete locations throughout cytoplasm
to perinuclear region that contains its
activator Rheb
The physical dissociation of mTORC1 and Rheb with amino acid
deprivation might explain why activators of Rheb (eg, growth factors) cannot
stimulate mTORC1 signalling in the absence of amino acids




mTORC2

 Knocking down mTORC2 components affects actin polymerisation and perturbs cell
morphology (Jacinto et al., 2004)
 Suggests mTORC2 controls actin cytoskeleton by promoting protein kinase (PKCα)
phosphorylation
 Signalling pathways that lead to mTORC2 activation not well determined – growth
factors increase mTORC2 kinase pathway and AKT phosphorylation at Ser473
considered plausible pathways

S6 kinase signalling

 S6 kinase is a central component implicated in growth signalling pathways
 Using S6K knockout animals display a small phenotype – only cell size affected, not
cell number
 Signalling pathways activated by growth factors
and nutrients converge on S6K, resulting in
kinase activation and up-regulation of cell growth
 Growth factors act via PI3K pathway
 Nutrients/energy status by the mTOR pathway
 Uncontrolled cell growth can occur via oncogenic
activation – particular genes within pathways
shown to be mutated or expressed at unusually
high levels, causing up-regulation of these
pathways
 S6K is a serine/threonine kinase from AGC family of kinases
- Number of identified substrates – when phosphorylated affect a number of
cellular processes, translation, RNA processing, survival and transcription