Summary for mammalian circadian biology: elucidating genome-wide levels of
• Circadian system – 1. SCN – in hypothalamus—entrained by daily light / dark
cycle, 2. Transmit synchronizing signal to local circadian oscillates in peripheral
• Biology clock/circadian rhythm – found in eukarya and bacteria, and some
members of archaea.
• Why need clock? – ie. cyanobacteria—oxygenic photosynthesis and nitrogen
fixation—need to be separated either temporally or spatially.
• a timing system, which regardless of environmental pressures, can create an
internal biological day synchronized to the external world enables an organism
to predict and prepare for daily environmental fluctuations
• Konopka and Benzer – molecular mechanism of the circadian clock consists of
a set of core clock genes and their protein products
• Orthologous—relate to the ancestor—some organism that diverged from a
common ancestor million yrs ago—many circadian clock genes are
Overview of the mammalian circadian system
• 3 major components of circadian systems:
o Light input pathway to a self sustained master circadian packemaker–
ganglion cell containing photo pigment, melanopsin
rod-cone system + melanopsin-containing ganglion cells—
serve visual function and entrainment of the circadian system
photic information to retina to hypothalamus via
RHT(retinalhypothalimic tract) to SCN (location of master
circadian pacemaker, located in the anterior hypothalamus)
o the circadian pacemaker
SCN contains about 8000-10000 neurons, which maintain
circadian cycle detached from SCN
SCN neurons express synchronized circadian rhythms of
spontaneous electrical activity, calcium oscillations, humoral
ouptput, metabolic activity, and gene expression
o output pathways by which the circadian pacemaker regulates overt
rhythms in biochemistry, physiology and behaviour throughout the
light enters SCN neural and hormonal out put signals that
influence various rhythms in the body
SCN regulates sleep/wake via intrahypothalamic projections
SCN signals impringe onANS
TGFa and prokineticin surpresses locomotor activity Cicadian phenotypes: Wheel running locomotor activity
• Zeitgeber times (ZT) (time giver) == any daily environmental cure to which –
circadian sysmter can synchronize and entrain.—i.e. light, may also be
temperature and social cue.
• When light deprived: Light pulse at –
o early night -maximum phase delay
late night - maximum phase advanced
o at day - small phase shift or none at all
The Mammalian core oscillatory mechanism
• core circadian clock genes code for proteins which form the primary molecular
circadian oscillatory mechanism
• Common feature of different species—participation of core clock gene in
interconnected positive /negative auto regulatory feed back loops of
transcription + translation.
• circadian rhythms are genetically based arising from cycles of transcription,
translation, protein-protein interaction, phosphorylation, nuclear translocation,
and protein degradation all impose delays at various times every day to
created coordinated 24h cycle
• Mammal – two core clock gene—1. Clock (express in SCN constitutively), 2.
Bmal1—peaks at middle of night.
• Clock/Bmal1 heterodimerize (CLOCK:BMAL1 cis or M34) – activate
transcription at E-box cis-regulatory enhancer sequence.
• Clock/Bmal1 complex—activate core gene 1. Per—PAS protein family
(Per1, 2, 3), 2. Cry – photoreceptor/photolyse family (Cry 1, 2)– peak as
opposite as Bmal1 peak so mid to late day since BMAL1 is peak during mid
• Core clock gene – after delayed by transcription—Per/Cry complex—go to
nucleus—stop Clock/Bmal complex—lower Per/Cry level directly—mainly
cause by Cry.
• Positive feedback – daily transcriptional activity of CLOCK:BMAL1
• Negative feedback – stopping CLOCK:BMAL1 transcription by CRYs to
inhibit their own transcription
• Rev-erbα—activated by Clock/Bmal1 complex—acting on E-box enhacer.
• Rev-erbaα—1. Indirectly inhibit own transcription by repressing at least
one of its activator Bmal1, by puttinn RORES in its promoter sequence,
whereas CRYAND PER associate with CLOCKAND BMAL directly, 2.
Also inhibited by repression of Cry/per complex on CLOCK: BMAL1.
• Inhibition of Rev-erba== timed circadian nuclear accumulation of
Clock/Bmal1—mainly Bmal1 dependent—activation of Bmal1 depend on
• Phosphorylation of core circadian protein—function degradation – regulated by casein kinase I epsilon (CKIE)—thus form 24-hr rhythms of RNA/protein level.
Mutations in Mammalian Core Clock Genes
• Clock gene mutation lengthens the circadian period
• BmalI/Mop3 mutation causes complete loss of circadian rhythmicity in
• of Per y
• Mutant Cry—1. Cry1 null mutant – 1 hr shorter , 2. Cry2 mutant – 1 hr
longer, 3. lose both—arrythmicity.
• Mutant Per—double mutant of Per1/Per2—strong circadian phenotype—if
only Per3—slight shorten circadian period— may play a role in clock output
• Syrian hamster tau mutation—in the gene coding CKIE?—1 mammal single
gene circadian mutation—result of C-T transition—proof—1. SCN master
circadian pacemaker, 2.Adiffusible signal can drive circadian rhythms in
animal, 3. SCN-independent circadian oscillator exists in mammal retina.
• Rev-erbα knockout—shorten free-running behavioral rhythms
Time now for Timeless?
• TIM/PER in Drosophila fly—same as CRY/PER complex in mammal –
translocate to nucleus inhibit