The basic-helix–loop–helix-PAS orphan MOP3 forms transcriptionally active complexes with circadian and hypoxia factors

Abstract

We report that MOP3 is a general dimerization partner for a subset of the basic-helix-loop-helix (bHLH) The PAS domain is found in a variety of proteins that play roles in development and adaptation to the environment (1-6). The PAS domain is also commonly found in proteins that harbor basic-helix-loop-helix (bHLH) domains, and that act in pairs as heterodimeric transcription factors (6-8). For example, the aryl hydrocarbon receptor nuclear translocator (ARNT) protein has been shown to act as a general partner for a number structurally related proteins that appear to act as sensors for environmental stimuli. ARNT dimerizes with the aryl hydrocarbon receptor (AHR) and mediates metabolic responses to 2,3,7,8-tetrachlorodibenzo-p-dioxin and related environmental contaminants. ARNT also dimerizes with hypoxiainducible factor 1␣ (HIF1␣) and mediates cellular responses to low oxygen and glucose (2, 6).-Recently, a number of ''orphan'' bHLH-PAS proteins have emerged from searches of expressed sequence tag databases and low stringency hybridization screens (9-14). For newly discovered bHLH-PAS proteins that have close homologs (e.g., HIF1␣ and HIF2␣ § or ARNT and ARNT2), partnering and DNA binding specificity can often be predicted from amino acid sequence similarities in their bHLH-PAS domains (9,12,14). For divergent orphans like MOP3, MOP4, and MOP5, amino acid sequence does not provide the information necessary for similar predictions. In an attempt to characterize this class of orphans, we have employed a series of assays that allow us to: (i) identify heterodimeric partnerships, (ii) determine the DNA response element bound by these heterodimers, (iii) verify that these complexes drive transcription in mammalian cells, and (iv) identify those tissues where these partnerships may occur. This report describes application of this approach to two bHLH-PAS orphans, MOP3 and MOP4.

The role of clock genes and rhythmicity in the liver

“…The hepatic oscillator is centered on a pair of transcriptional activators, BMAL1 (also named Mop3) (Bunger et al, 2000;Hogenesch et al, 1998) and CLOCK (Gekakis et al, 1998;King et al, 1997) and two classes of repressors of the Period (Per) Shearman et al, 1997;Sun et al, 1997;Tei et al, 1997) and Cryptochrome (Cry) gene families (Griffin et al, 1999;Kume et al, 1999;van der Horst et al, 1999;Vitaterna et al, 1999) (Fig. 2).…”

Assessment of circadian function in fibroblasts of patients with bipolar disorder

“…39,40 The positive feedback loop is composed of CLOCK and BMAL1 heterodimer which activates Period (PER1, PER2 and PER3) and Cryptochrome (CRY1 and CRY2) transcription. [41][42][43][44] The negative feedback loop involves rhythmic inhibition of the transcriptional activity of CLOCK/BMAL1 by the PER and CRY complexes. [45][46][47][48] These negative regulators also suppress the expression of REV-ERBa which inhibits BMAL1 transcription through RORE elements in the BMAL1 promoter.…”

Intermolecular recognition revealed by the complex structure of human CLOCK-BMAL1 basic helix-loop-helix domains with E-box DNA

“…The E-box is distinguished by having a consensus sequence of CANNTG [13][14][15], with a palindromic canonical form of CACGTG [16]. By site selection and amplification method, the MOP4-BMAL1 (MOP4, also named NPAS2, a homolog of CLOCK) heterodimer was demonstrated to bind the canonical E-box, which is also considered to be a high-affinity binding site for CLOCK-BMAL1 [7]. Further large-scale studies on cycling genes in suprachiasmatic nuclei, the liver and the heart have shown that a number of non-canonical E-box elements also regulate rhythmic gene expression [17][18][19].…”

“…The transcription control of the core loop is governed by two bHLH (basic helix-loop-helix) and PAS (period-ARNTsingle-minded) domain-containing transcription factors, namely CLOCK (circadian locomotor output cycles kaput) and BMAL1 (brain and muscle ARNT-like 1) [1,2,5,6]. It has been established that they form a heterodimer, bind E-box DNA elements and activate the transcription of clock-controlled genes [7,8]. Two groups of resultant proteins, PERIOD (PER, sum of PER1-3) and CRYPTO-CHROME (CRY, or CRY1, 2), gradually accumulate and inhibit the activity of CLOCK-BMAL1 complex, constituting the negative feedback limb of the core loop [1,2,5,6].…”