Molecular Playground/CLOCK:BMAL1 heterodimer complex

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CLOCK:BMAL1 heterodimer complex

The is a vital regulatory component of the circadian rhythm protein regulation system. CLOCK (Circadian Locomotor Output Cycles Kaput) and BMAL1 (Brain and muscle Arnt-like protein-1) are PER-ARNT-SIM (bHLH-PAS) proteins. (The structure highlighted in yellow is a portion of bHLH structure in CLOCK) The CLOCK:BMAL1 heterodimer is the main transcriptional activator in the mammalian circadian mechanism.^[1] The binding between CLOCK and BMAL1 involves the N-terminal bHLH, PAS-A and PAS-B domains of both proteins. Each domain (bHLH, PAS-A, PAS-B) binds to its corresponding equivalent in the other protein. Though both proteins contain the same types of domains with similar primary amino acid sequences in each, the overall heterodimer is surprisingly asymmetrical due to differences in the spatial orientation of the domains in each protein. Since this heterodimer complex involves the binding of all of the major domains in both participating proteins, the overall binding affinity is very high.

1 Role in Circadian Rhythm
2 The Overall structure of CLOCK:BMAL1 complex
- 2.1 CLOCK
- 2.2 BMAL1
3 The interface between CLOCK and BMAL1
4 The downstream effects of the altered circadian rhythm

Role in Circadian Rhythm

Circadian rhythms are operated by an endogenous core clock system that drives daily rhythms in behavior, physiology, and metabolism. In mammalian systems, the suprachiasmatic nucleus (SCN), which is located in the hypothalamus, is the locus of a master circadian clock. The SCN controls the expression of of proteins in a time dependent manner through a genetic feedback loop initiated by light passing through the eye.^[2] The core molecular clockwork is composed of a transcriptional/post-translational feedback loop: CLOCK:BMAL1 (transcriptional activators) and PER:CRY (transcriptional repressors). In daytime, CLOCK and BMAL1 will form a heterodimer complex to bind the E-box promoter region of other circadian rhythm proteins, causing the transcription of Per (Period) and Cry (Cryptochrome). During the day, Per and Cry will dimerize and translocate into the nucleus, where they interact with CLOCK:BMAL1 to inhibit their own transcription, forming a negative feedback loop.^[3] At night time, Per:Cry complex is degraded by a specific E3 ligase complex and the repression is relieved. After the repression level of Per:Cry is decreased, CLOCK:BMAL1 will be re-activated and start transcription again, forming the positive feedback loop. The entire negative/positive feedback loop take around 24 hours to complete, thus forming the core mechanism of the circadian clock in mammals.^[4]

The Overall structure of CLOCK:BMAL1 complex

The is shown in the right. It is a tightly intertwined structure where CLOCK and BMAL1 are twisted together. Although the primary sequences of CLOCK and BMAL1 are similar, the structural arrangements of their domains are quite different.

CLOCK

is composed of three domains: one domain, two PAS domains ( and ). The connections between each domain are two . Comparedto the flexible loops in BMAL1, the distances of the connection loops in CLOCK are longer.

BMAL1

is also composed of three domains: one domain, two PAS domains ( and ). There is a ~15-residue flexible loop () and a ~20-residue flexible loop () connecting between each domain.

The interface between CLOCK and BMAL1

In the formation of the CLOCK:BMAL1 heterodimer complex, each domain interacts with the corresponding domain of its partner subunit, i.e. CLOCK bHLH interacts with BMAL1 bHLH domain and so forth. In PSA domains, contain a five-stranded antiparallel β sheet and several α helices flanking the concave surface of the sheet. In those α helices, there are two packed in between the β-sheet faces of the two domains to mediate the heterodimeric PSA-A interactions.

PSA-A domain interface

The interface bin the CLOCK:BMAL1 PSA-A dimer is mainly facilitated by hydrophobic interactions. Specifically, Phe104, Leu105, and Leu113 on (Leu159, Thr285, Tyr287, Val315, and Ile317). Similarly, Phe141, Leu142, and Leu150 on BMAL1 PSA-A contact the hydrophobic β-sheet face of CLOCK (F122, I216, V252, and T254). As a result, many residues obtained in the CLOCK:BMAL1 interface are conserved among bHLH-PAS transcription factors. This result may indicate that CLOCK and BMAL1 have a common PSA-A domain dimerization mode.

PSA-B domain interface

The PSA-B domains of CLOCK and BMAL1 are stacked in parallel conformation. The β-sheet on PSA-B domain of BMAL1 contact the helical face of CLOCK PSA-B domain. The bury some hydrophobic residues on both subunits, including Try310, Val315, and Leu318 of CLOCK and Phe423, Trp427, and Val435 of BMAL1.

The binding interface between CLOCK:BMAL1 and E-box element

Fig. 1 The overlap structure of CLOCK:BMAL1 complex with bHLH Myc:Max-DNA complex (pdb: 1NKP). The figure is generated by Pymol

As shown in Fig. 1, the four-helical bHLH bundle in the CLOCK:BMAL1 heterodimer overlaps with the bHLH Myc:Max-DNA complex (pdb: 1NKP).

The downstream effects of the altered circadian rhythm

It has been shown in recent years that people who have lifestyles which involve light exposure that is different than the normal 12 hours of daylight/12 hours of night have significantly increased chances of developing cancer. This indicates that disruption of the normal circadian rhythm gene regulation cycle has severe downstream effects on the host's genetic makeup. Additionally, it has been shown that cancer tissues often have distorted circadian rhythms, showing the significance of circadian rhythms to cancer progression. ^[5]

Mutated forms of CLOCK exist which do not regulate protein expression correctly and thereby result in altered circadian rhythms. CLOCK-delta19 is a mutant form of CLOCK which binds to BMAL1 normally but the resulting heterodimer does not activate transcription of certain other circadian rhythm proteins. Mutant mice carrying this altered CLOCK protein have shown abnormal circadian rhythms as a result. An example of the downstream effect of this mutant form of CLOCK is the resulting abnormal expression of NAMPT. In normal mice, NAMPT is expressed in a circadian manner, showing oscillations in expression regardless of light conditions. However, in mice with mutant CLOCK-delta19, NAMPT is not expressed in an circadian manner and the overall expression is lower. This leads to further issues as NAMPT is the rate limiting enzyme for the biosynthesis of NAD+, which is an important biological coenzyme.^[6]

References

↑ doi: https://dx.doi.org/10.1126/science.280.5369.1564
↑ Silver R, Kriegsfeld LJ. Circadian rhythms have broad implications for understanding brain and behavior. Eur J Neurosci. 2014 Jun;39(11):1866-80. doi: 10.1111/ejn.12593. Epub 2014 May 5. PMID:24799154 doi:http://dx.doi.org/10.1111/ejn.12593
↑ Huang N, Chelliah Y, Shan Y, Taylor CA, Yoo SH, Partch C, Green CB, Zhang H, Takahashi JS. Crystal structure of the heterodimeric CLOCK:BMAL1 transcriptional activator complex. Science. 2012 Jul 13;337(6091):189-94. Epub 2012 May 31. PMID:22653727 doi:10.1126/science.1222804
↑ Lowrey PL, Takahashi JS. Genetics of circadian rhythms in Mammalian model organisms. Adv Genet. 2011;74:175-230. doi: 10.1016/B978-0-12-387690-4.00006-4. PMID:21924978 doi:http://dx.doi.org/10.1016/B978-0-12-387690-4.00006-4
↑ Stevens RG. Circadian disruption and breast cancer: from melatonin to clock genes. Epidemiology. 2005 Mar;16(2):254-8. doi: 10.1097/01.ede.0000152525.21924.54. PMID:15703542 doi:http://dx.doi.org/10.1097/01.ede.0000152525.21924.54
↑ Ramsey KM, Yoshino J, Brace CS, Abrassart D, Kobayashi Y, Marcheva B, Hong HK, Chong JL, Buhr ED, Lee C, Takahashi JS, Imai S, Bass J. Circadian clock feedback cycle through NAMPT-mediated NAD+ biosynthesis. Science. 2009 May 1;324(5927):651-4. doi: 10.1126/science.1171641. Epub 2009 Mar , 19. PMID:19299583 doi:http://dx.doi.org/10.1126/science.1171641

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Retrieved from "http://52.214.119.220/wiki/index.php/Molecular_Playground/CLOCK:BMAL1_heterodimer_complex"

@@ Line 7: / Line 7: @@
 == Role in Circadian Rhythm ==
-Circadian rhythms are operated by an endogenous core clock system that drives daily rhythms in behavior, physiology, and metabolism. In mammalian systems, the suprachiasmatic nucleus (SCN), which is located in the hypothalamus, is the locus of a master circadian clock. The SCN controls the expression of of proteins in a time dependent manner through a genetic feedback loop initiated by light passing through the eye.<ref>doi: 10.1111/ejn.12593</ref> The core molecular clockwork is composed of a transcriptional/post-translational feedback loop: CLOCK:BMAL1 (transcriptional activators) and PER:CRY (transcriptional repressors). In daytime, CLOCK and BMAL1 will form a heterodimer complex to bind the E-box promoter region of other circadian rhythm proteins, causing the transcription of Per (''Period'') and Cry (''Cryptochrome''). During the day, Per and Cry will dimerize and translocate into the nucleus, where they interact with CLOCK:BMAL1 to inhibit their own transcription, forming a negative feedback loop.<ref>DOI: 10.1126/science.1222804</ref> At night time, Per:Cry complex is degraded by a specific E3 ligase complex and the repression is relieved. After the repression level of Per:Cry is decreased, CLOCK:BMAL1 will be re-activated and start transcription again. This process is called the positive feedback loop. The whole negative/positive feedback loops take around 24 h to complete, thus form the core mechanism of the circadian clock in mammals.<ref>doi:10.1016/B978-0-12-387690-4.00006-4</ref>
+Circadian rhythms are operated by an endogenous core clock system that drives daily rhythms in behavior, physiology, and metabolism. In mammalian systems, the suprachiasmatic nucleus (SCN), which is located in the hypothalamus, is the locus of a master circadian clock. The SCN controls the expression of of proteins in a time dependent manner through a genetic feedback loop initiated by light passing through the eye.<ref>doi: 10.1111/ejn.12593</ref> The core molecular clockwork is composed of a transcriptional/post-translational feedback loop: CLOCK:BMAL1 (transcriptional activators) and PER:CRY (transcriptional repressors). In daytime, CLOCK and BMAL1 will form a heterodimer complex to bind the E-box promoter region of other circadian rhythm proteins, causing the transcription of Per (''Period'') and Cry (''Cryptochrome''). During the day, Per and Cry will dimerize and translocate into the nucleus, where they interact with CLOCK:BMAL1 to inhibit their own transcription, forming a negative feedback loop.<ref>DOI: 10.1126/science.1222804</ref> At night time, Per:Cry complex is degraded by a specific E3 ligase complex and the repression is relieved. After the repression level of Per:Cry is decreased, CLOCK:BMAL1 will be re-activated and start transcription again, forming the positive feedback loop. The entire negative/positive feedback loop take around 24 hours to complete, thus forming the core mechanism of the circadian clock in mammals.<ref>doi:10.1016/B978-0-12-387690-4.00006-4</ref>
 ==The Overall structure of CLOCK:BMAL1 complex==
-The <scene name='60/609802/Clock_bmal1/1'>3D structure of CLOCK:BMAL1 heterodimer</scene> is shown in the right. It is a tightly interwined structure that CLOCK and BMAL1 are twisted together. Although the primary sequences of CLOCK and BMAL1 are similar, the structural arrangements of their domains are quite different.
+The <scene name='60/609802/Clock_bmal1/1'>3D structure of CLOCK:BMAL1 heterodimer</scene> is shown in the right. It is a tightly intertwined structure where CLOCK and BMAL1 are twisted together. Although the primary sequences of CLOCK and BMAL1 are similar, the structural arrangements of their domains are quite different.
 ===CLOCK===
-<scene name='60/609802/Clock_only/1'>CLOCK</scene> is composed of three domains: one <scene name='60/609802/Clock_bhlh/1'>N-terminal bHLH</scene> domain, two PAS domains (<scene name='60/609802/Clock_psa_a/1'>PSA-A</scene> and <scene name='60/609802/Clock_psa_b/1'>PSA-B</scene>). The connections between each domain are two <scene name='60/609802/Clock_l1_l2/1'>flexible loops</scene>. Comparing to the flexible loops in BMAL1, the distances of the connection loops in CLOCK are longer.
+<scene name='60/609802/Clock_only/1'>CLOCK</scene> is composed of three domains: one <scene name='60/609802/Clock_bhlh/1'>N-terminal bHLH</scene> domain, two PAS domains (<scene name='60/609802/Clock_psa_a/1'>PSA-A</scene> and <scene name='60/609802/Clock_psa_b/1'>PSA-B</scene>). The connections between each domain are two <scene name='60/609802/Clock_l1_l2/1'>flexible loops</scene>. Comparedto the flexible loops in BMAL1, the distances of the connection loops in CLOCK are longer.
 ===BMAL1===
-<scene name='60/609802/Bmal1_only/1'>BMAL1</scene> is also composed of three domains: one <scene name='60/609802/Bmal1_bhlh/1'>N-terminal bHLH</scene> domain, two PAS domains (<scene name='60/609802/Bmal1_psa_a/1'>PSA-A</scene> and <scene name='60/609802/Bmal1_psa_b/1'>PSA-B</scene>). There are ~15-residue flexible loop (<scene name='60/609802/Bmal1_l1/1'>L1</scene>) and ~20-residue flexible loop (<scene name='60/609802/Bmal1_l2/1'>L2</scene>) connect between each domains.
+<scene name='60/609802/Bmal1_only/1'>BMAL1</scene> is also composed of three domains: one <scene name='60/609802/Bmal1_bhlh/1'>N-terminal bHLH</scene> domain, two PAS domains (<scene name='60/609802/Bmal1_psa_a/1'>PSA-A</scene> and <scene name='60/609802/Bmal1_psa_b/1'>PSA-B</scene>). There is a ~15-residue flexible loop (<scene name='60/609802/Bmal1_l1/1'>L1</scene>) and a ~20-residue flexible loop (<scene name='60/609802/Bmal1_l2/1'>L2</scene>) connecting between each domain.
 ==The interface between CLOCK and BMAL1==
-In the formation of CLOCK:BMAL1 heterodimer complex, each domain interacts with the corresponding domain of its partner subunit, which means CLOCK bHLH interacts with BMAL1 bHLH domain, CLOCK PSA-A interacts with BMAL1 PSA-A domain, and CLOCK PSA-B interacts with BMAL1 PSA-B domain. In PSA domains, <scene name='60/609802/Clock_bmal1_psa_a/1'>both CLOCK and BMAL1 PSA-A domains</scene> contain five-stranded antiparallel β sheet and several α helices flanking the concave surface of the sheet. In those α helices, there are two <scene name='60/609802/Clock_bmal1_a_alpha/1'>N-terminal flanking helix (A'α) externals</scene> pack in between the β-sheet faces of two domains to mediate the heterodimeric PSA-A interactions.
+In the formation of the CLOCK:BMAL1 heterodimer complex, each domain interacts with the corresponding domain of its partner subunit, i.e. CLOCK bHLH interacts with BMAL1 bHLH domain and so forth. In PSA domains, <scene name='60/609802/Clock_bmal1_psa_a/1'>both CLOCK and BMAL1 PSA-A domains</scene> contain a five-stranded antiparallel β sheet and several α helices flanking the concave surface of the sheet. In those α helices, there are two <scene name='60/609802/Clock_bmal1_a_alpha/1'>N-terminal flanking helices (A'α) externals</scene> packed in between the β-sheet faces of the two domains to mediate the heterodimeric PSA-A interactions.
 === PSA-A domain interface ===
-The interface between CLOCK:BMAL1 PSA-A dimer is mainly facilitated by hydrophobic interactions. Specifically, Phe104, Leu105, and Leu113 on <scene name='60/609802/Clock_bmal1_if_1/1'>the A'α helix of CLOCK contact the hydrophobic region on the β-sheet face of BMAL1</scene> (Leu159, Thr285, Tyr287, Val315, and Ile317). Similarly, Phe141, Leu142, and Leu150 on BMAL1 PSA-A contact to the hydrophobic β-sheet face of CLOCK (F122, I216, V252, and T254). As a result, many residues obtained in the CLOCK:BMAL1 interface are conserved among bHLH-PAS transcription factors. This result may indicate that CLOCK and BMAL1 have a common PSA-A domain dimerization mode.
+The interface bin the CLOCK:BMAL1 PSA-A dimer is mainly facilitated by hydrophobic interactions. Specifically, Phe104, Leu105, and Leu113 on <scene name='60/609802/Clock_bmal1_if_1/1'>the A'α helix of CLOCK contact the hydrophobic region on the β-sheet face of BMAL1</scene> (Leu159, Thr285, Tyr287, Val315, and Ile317). Similarly, Phe141, Leu142, and Leu150 on BMAL1 PSA-A contact the hydrophobic β-sheet face of CLOCK (F122, I216, V252, and T254). As a result, many residues obtained in the CLOCK:BMAL1 interface are conserved among bHLH-PAS transcription factors. This result may indicate that CLOCK and BMAL1 have a common PSA-A domain dimerization mode.
 ===PSA-B domain interface===
-The PSA-B domains of CLOCK and BMAL1 are stacked in parallel conformation. The β-sheet on PSA-B domain of BMAL1 contacts with the helical face of CLOCK PSA-B domain. The <scene name='60/609802/Clock_bmal1_if_2/1'>contact interface</scene> bury some hydrophobic residues on both subunits, including Try310, Val315, and Leu318 of CLOCK and Phe423, Trp427, and Val435 of BMAL1.
+The PSA-B domains of CLOCK and BMAL1 are stacked in parallel conformation. The β-sheet on PSA-B domain of BMAL1 contact the helical face of CLOCK PSA-B domain. The <scene name='60/609802/Clock_bmal1_if_2/1'>contact interface</scene> bury some hydrophobic residues on both subunits, including Try310, Val315, and Leu318 of CLOCK and Phe423, Trp427, and Val435 of BMAL1.
 ===The binding interface between CLOCK:BMAL1 and E-box element===