Hormonal Regulation (Lecture 33) Flashcards
Metabolic regulation in complex organisms
Short term regulation (seconds, minutes) = achieved through allosteric control (ie. level
of metabolites) and post-translational modifications (ie. phosphorylation) of enzymes
under hormonal control (eg. catecholamines, insulin)
- Long term regulation (hours, circadian cycle, seasons, development, aging, etc.) =
requires transcriptional regulation of metabolic genes
Transcriptional Control of Metabolism
Requires specific signals to be transduced to nucleus where individual genes or entire gene
networks are targeted for regulation
Important Aspects of transcriptional control of specific metabolic responses:
- Events upstream of transcriptional activity => signals involved (eg. glucagon, glucocorticoids) and their route to the nucleus (eg. signalling pathways, protein cleavage, direct activation)
- Molecular mechanisms by which transcription factors regulate gene expression
Eg. recruitment of polymerase, enzymatic activity of coregulators - Events downstream of transcription => depend on genes being targeted and which further signals are generated
Eg. expression of metabolic enzymes or cascade of regulators
Metabolic Transcription Factors
Bind DNA and receive signals to tell TFs to activate/repress transcription
Nuclear Receptors
family of 48 ligand-responsive “zinc fingers” transcription factors
- Many members work as a metabolic sensor involved in all aspects of metabolism
(carbohydrate, lipid, amino acid, etc.) - Most NRs are directly activated by their ligands => hormones will enter nucleus from outside of cell and directly bind receptor
- Can also be regulated by post-translational modifications
CREB (cAMP Response Element Binding)
CREM and ATFI
“leucine zipper” TF activated by phosphorylation by PKA in response to increased cAMP levels following glucagon action
- Considered first responders in activation of gluconeogenesis during fasting
CEBP (CCAAT-enhancer binding protein)
basic “leucine zipper” TF
- Activity is constitutive
- Signal-independent high expression in liver plays major role in response to
fasting
Forkhead Box Proteins
FoxO1, FoxO3, FoxO4, FoxO6, FoxA2)
involved in hepatic (liver) glucose production after nutrient deprivation
- Factors are activated by post-translational modifications such as phosphorylation and acetylation
SREBP-1c (sterol response element binding protein)
bHLH-leucine zipper TF that targets genes involved in lipid metabolism
- Involved in cholesterol synthesis
- It is activated by proteolytic cleavage stimulated by sterols and unsaturated FAs
=> dormant in ER, low sterol levels activate protein which is trafficked to Golgi, recruties protease that cleaves twice and sends small part to nucleus for transcriptional activation of genes involved in sterol synthesis
hREBP (carbohydrate response element binding protein)
bHLH-leucine zipper TF phosphorylated by PKA
- Responsive to glucose
- Phosphorylated ChREBP via PKA= inactive ChREBP
- Activated by removal of phosphate groups by PP2A phosphatase
- Regulates glucose and lipid metabolism
Metabolic Coregulators
- Do not bind DNA, interact with TFs and or other coregulators
- TF binding proteins recognize and bind DNA sequence but coregulators are the moiety which activate or repress gene expression
NCOA
nuclear receptor coactivator
Aka SRC, steroid receptor coactivator
Family of 3 members involved in all aspects of metabolism (lipid, carbohydrate, aa)
NCOR 1 and 2
nuclear receptor corepressor
Two corepressors that often oppose action of NCOAs and other co-activator proteins
RIP140 = aka NRIP1
- Action as repressor decreases mitochondrial biogenesis and oxidative metabolism in muscle
- Especially at rest
PGC-1alpha and beta
master co-activators that interact with multiple transcription factors, most predominantly the nuclear receptors PPARs and ERRs
- Required for mitochondrial biogenesis, oxidative metabolism and heat production by brown fat
KAT2A and B
lysine acetyltransferase Aka GCN5 and PCAF, which are histone acetyltransferases
- KAT2alpha acetylated PGC-1alpha to decrease its activity and blunting PGC-1alpha
induced gluconeogenesis
HDACs
large family of histone deacetylases with broad roles in metabolic control
- Recruitment of HDAC3 by nuclear receptor Rev-Erb-alpha (clock gene) inhibits expression of lipogenic genes
CRTCs
family of 3 proteins that act as coactivators of CREB
Regulate CREB-dependent hepatic gluconeogenic program
PRMTI
methylates nuclear receptors,
- in particular HNF-4alpha
SRTI
a sirtuin, NAD+-dependent deacetylase that targets PGC-1alpha, ERRalpha and other coactivators
Affect mitochondrial function
Nuclear Receptors
Are regulators of metabolism
Protypic TF to study gene regulation
Clinically relevant =>Targets of several drugs to control metabolism
Some are in cytoplasm, bind hormones and move to nucleus
Others are already bound to DNA in nucleus, awaiting hormonal signalling
Recruit coactivators which change expression of target genes
Transcribe genes that will affect metabolic activity
Nuclear receptors and other TFs do not have enzymatic activity; their function is to
recruit co regulatory proteins on DNA to activity transcription
what are the ligands for nuclear receptors
hormonal ligans and metabolic ligands
DHT
male steroid hormone, anabolic steroid for building muscle/strength
Estrogen
female steroid hormone, involved in metabolic pathway, preparing for development of embryo, fat metabolism, change in metabolism associated with estrogen decrease at menopause often leads to weight gain
cortisol
corticosteroid expressed in all tissues and regulate glucose
metabolism via glucocorticoid receptor
Thyroid hormone
important in regulating metabolism; hyperthyroidism => high
metabolism, lose weight, opposite can also happen
3 main elements of nuclear receptors
DNA. binding domain
modulator
ligand binding domain
modulator function
the modulator is located at the N terminus and functions to modulate receptor activity. this is the target for phosphorylation and it receives signals through phosphorylation and acetylation. a trans activation through coregualttion interaction (AF-1) also occurs at this site
DNA binding domain
DNA binding domain recognizes a specific sequence of DNA
it is composed of zinc fingers and requires dimerization
ligand binding domain
receives signals from ligands when bound to the nuclear receptor
can form homo and hetero dimers
transactivation (AF-2) where the coavtivators bind to the receptors for coregualtion interaction
Nuclear receptors recognize specific DNA sequences near metabolic genes
Nuclear receptors recognizes hormone response element (HRE)
methods of recognizing the HRE
Monomer can recognize extended half site
Dimers can recognize two sites
Homodimers act on inverted repeats
Heterodimers act on direct repeats
core motifs
AGGTCA (TGCCT)
AGAACA (TGTTCT)
Genome-wide identification of target genes
Must identify target genes of receptors
Stimuli activate certain receptors => can observe mitochondrial
biogenesis, fatty acid oxidation, oxphos, etc.
when suing ChIP- sequencing (chromatic immunoprecipitation) to identify genome wide targets, the following stops are carried out:
(1) Extraction of DNA with factor bound
(2) Fix factor to DNA with formaldehyde
(3) Cut into small pieces
(4) Use antibody against factor of interest, linked to beads
(5) Immunoprecipitate beads, beads often magnetic
(6) Add linkers to DNA, amplify by PCR, sequence & analyze
what can be determined with ERR-alpha
the identification of metabolic pathways
allows for the generation of a map of activators and inhibitors
cistrome
entire set of binding events for a given transcription factor
ChIP sequencing identifies
target genes of transcription factors and their cistrome
also shows the binding spots, regulatory regions near metabolic genes that contains sites for TFs
this indicated that transcriptional regulation of metabolic genes requires the coordination between several factors
how do Nuclear Receptors require coregulators to modulate gene expression?
Nuclear receptors act as activators and repressors of metabolic genes
Switch between corepressors and coactivator complexes allows TFs to act both negatively and positively on their targets in response to specific extracellular and intracellular signals
Intracellular = metabolite levels Extracellular = hormones
Factors involved in exchange of corepressor/activator complexes as well as TF are targets of multiple extracellular and intracellular pathways
