Lecture 4- Genetic components of the mammalian molecular clock Flashcards
DBP
“classic” clock output gene, rhythmic expression of which was identified by Shibler (150x stronger in day vs night)
Pleiotropic effects of clocks
NB pleiotropy is where one gene has one or more seemingly unrelated phenotypic effects
sleep/ wake, cell cycle progression, DNA damage , metabolism, energy metabolism, immune responses
How much of the genome is under circadian control?
10%
Disruptions in clock genes are related to..
Sleep disorders, mental health problems, obesity, cancer CV problems
What makes the SCN unique?
The only somatic clock that receives light input (NB neurons types in the SCN are not unique)
How many cells are in the SCN?
20,000
SCN output
behavioural, hormonal, peripheral opscillators
Examples of cells that do not possess clocks?
Embryonic stem cells and germline cells
What happens to the cellular clock if a cell reverts back to its embryonic state?
Cellular clocks are lost
Mutagenesis and genome wide screening
… have been to show that the basic building blocks of clocks are highly conserved between animals
Other than mutagenesis and GWS, what methods have been employed to find clock genes?
Behavioural studies in mice and flies
How was the gene PERIOD identified?
PERIOD1,2,3 identified by mutagenesis in Drosophila, then cloned by homology in mammals
How was the gene CLOCK identified?
Mutagenesis in mammals
How was the gene BMAL1 identified?
As an interacting protein (with CLOCK) in mammals
How was CRY identified?
CRY1,2 identified by clock relevant mutagenesis in Drosophila
How was Casein Kinase 1 ε (CK1ε) identified?
As a spontaneous tau mutation (hamster)
CK1ε function
Binds to and phosphorylates PER proteins, both of which leads to their degradation and prevents them from being able to enter the nucleus and suppress the transcription of clock genes
BMAL1 and CLOCK w/r/t PER and CRY
bind to the Ebox elements of the nuclear DNA, but PER and CRY do not, instead they bind directly to BMAL1 and CLOCK
Phosphorylation of PER and CRY…
… targets them to the 26s proteasome for degradation (as well as causing them to detach from BMAL1 and CLOCK)
Where are clock regulatory elements found?
In promoter regions of downstream genes
REV-ERB proteins
Nuclear receptors and transcriptional repressors of BMAL1
ROR proteins
Nuclear receptors and transcriptional activators of BMAL1
Mutations in CK1ε
Alter cycle length (HOW)
What are the core clock genes?
CLOCK, BMAL1, PER, CRY, REV-ERBalpha, ROR
PER KO studies
Gradual loss of rhythmicity in PER1/2 KOs but not in PER3 KOs; PERs are essential but PER3 is unlikely to be involved in locomotor behaviour, more likely sleep/wake regulation; paired PER1/3 and PER2/3 KOs have indicated partial redundancy between PER1 and PER2
CRY KO studies
in DD, KO of CRY1 or CRY2 = free running wherease a double KO = arrhythmic –> partial redundancy but are essential
CLOCK KO studies
CLOCK is non essential and mice w/o exhibit normal circadian rhythm
Which TF makes CLOCK partially redundant?
NPAS2
Only clock gene with no redundancy and is completely essential
BMAL1
Tau (C->T) mutation in CK1ε
More active degrader of PER2 –> speeds up clock by ~3hours
CK1delta inhibitors
increase the period of circadian clocks
Modulation of REV-ERBalpha
resets the circadian clock in a phasic manner
Lithium
increases amplitude and period of circadian clock
Non-pharma improvement of circadian rhythm
regular meal times, scheduled exercise, light therapy
