Biomedical subjects

C Vinson

Publications and source records attributed to C Vinson.

At least 37 records · Page 2Linked to original sources

PubMed1999

Torpor in mice is induced by both leptin-dependent and -independent mechanisms.

We tested the effect of chronic leptin treatment on fasting-induced torpor in leptin-deficient A-ZIP/F-1 and ob/ob mice. A-ZIP/F-1 mice have virtually no white adipose tissue and low leptin levels, whereas ob/ob mice have an abundance of fat but no leptin. These two models allowed us to examine the roles of adipose tissue and leptin in the regulation of entry into torpor. Torpor is a short-term hibernation-like state that allows conservation of metabolic fuels. We first characterized the A-ZIP/F-1 animals, which have a 10-fold reduction in total body triglyceride stores. Upon fasting, A-ZIP/F-1 mice develop a lower metabolic rate and decreased plasma glucose, insulin, and triglyceride levels, with no increase in free fatty acids or beta-hydroxybutyrate. Unlike control mice, by 24 hr of fasting, they have nearly exhausted their triglycerides and are catabolizing protein. To conserve energy supplies during fasting, A-ZIP/F-1 (but not control) mice entered deep torpor, with a minimum core body temperature of 24 degrees C, 2 degrees C above ambient. In ob/ob mice, fasting-induced torpor was completely reversed by leptin treatment. In contrast, neither leptin nor thyroid hormone prevented torpor in A-ZIP/F-1 mice. These data suggest that there are at least two signals for entry into torpor in mice, a low leptin level and another signal that is independent of leptin and thyroid hormone levels. Studying rodent torpor provides insight into human torpor-like states such as near drowning in cold water and induced hypothermia for surgery.

Adaptation, Physiological↗

PubMed1999

Transgenic mice lacking white fat: models for understanding human lipoatrophic diabetes.

The human disease lipoatrophic (or lipodystrophic) diabetes is a rare syndrome in which a deficiency of adipose tissue is associated with Type 2 diabetes. This disease is an interesting contrast to the usual situation in which diabetes is associated with obesity, an excess of fat. Aside from obesity, patients with lipodystrophic diabetes have the other features associated with Metabolic Syndrome X, including hypertension and dyslipidemia. The contrast between diabetes with a lack of fat and diabetes with an excess of fat provides an opportunity to study the mechanisms causing Type 2 diabetes and its complications. Recently, three laboratories have produced transgenic mice that are deficient in white adipose tissue. These mice have insulin resistance and other features of lipoatrophic diabetes, and are a faithful model for the human disease. Here we review the different murine models of fat ablation and compare the murine and human diseases, addressing the questions: Is the lack of fat causative of the diabetes, and if so by what mechanism? How could the other clinical features be explained mechanistically? And finally, what can be gleaned about insight into treatment options?

Animals↗

PubMed1999

Fibrin fragment induction of plasminogen activator inhibitor transcription is mediated by activator protein-1 through a highly conserved element.

Plasminogen activator inhibitor type-1 (PAI-1), a serine protease inhibitor, affects the processes of fibrinolysis, wound healing, and vascular remodeling. We have demonstrated that PAI-1 transcription is induced by D dimer, a plasmin proteolytic fragment of fibrin, supporting its role in negative feedback on peri-cellular proteolysis. The focus of this study was to define the mechanism of D dimer's effects on PAI-1 transcription. D dimer increased the binding activity of the transcription factor activator protein-1 components c-fos/junD and c-fos mRNA levels in a time- and concentration-dependent manner to a greater extent than fibrinogen. Both basal and D dimer-induced PAI-1 transcriptional activity were entirely dependent on elements within the -161 to -48 bp region of the PAI-1 gene in fibroblasts. Mutations within the AP-1-like element (-59 to -52 bp) in the PAI-1 gene affected D dimer-induced transcriptional activity, c-fos/junD DNA binding, and basal and c-fos inducible PAI-1 transcriptional activity. Furthermore, expression of either wild-type or mutant c-fos proteins augmented or diminished the response of the PAI-1 promoter (-161 to +26 bp) to D dimer, respectively. D dimer-induced binding of c-fos/junD to the highly conserved and unique AP-1 like element in the PAI-1 gene provides a mechanism whereby specific fibrin fragments control fibrin persistence at sites of inflammation, fibrosis, and neoplasia.

Animals↗

PubMed1999

Bacterial peptidoglycan induces CD14-dependent activation of transcription factors CREB/ATF and AP-1.

Peptidoglycan (PGN), the major cell wall component of Gram-positive bacteria, induces secretion of cytokines in macrophages through CD14, the pattern recognition receptor that binds lipopolysaccharide and other microbial products. To begin to elucidate the mechanisms that regulate the transcription of cytokine genes, we wanted to determine which transcription factors are activated by PGN in mouse RAW264.7 and human THP-1 macrophage cells. Our results demonstrated that: (i) PGN induced phosphorylation of the transcription factors ATF-1 and CREB; (ii) ATF-1 and CREB bound DNA as a dimer and induced transcriptional activation of a CRE reporter plasmid, which was inhibited by dominant negative CREB and ATF-1; (iii) PGN induced phosphorylation of c-Jun, protein synthesis of JunB and c-Fos, and transcriptional activation of the AP-1 reporter plasmid, which was inhibited by dominant negative c-Fos; and (iv) PGN-induced activation of CREB/ATF and AP-1 was mediated through CD14. This is the first study to demonstrate activation of CREB/ATF and AP-1 transcription factors by PGN or by any other component of Gram-positive bacteria.

Activating Transcription Factor 1↗

PubMed1999

CRE DNA binding proteins bind to the AP-1 target sequence and suppress AP-1 transcriptional activity in mouse keratinocytes.

Previously, we have shown that nuclear extracts from cultured mouse keratinocytes induced to differentiate by increasing the levels of extra-cellular calcium contain Fra-1, Fra-2, Jun B, Jun D and c-Jun proteins that bind to the AP-1 DNA binding sequence. Despite this DNA binding activity, AP-1 reporter activity was suppressed in these cells. Here, we have detected the CREB family proteins CREB and CREMalpha as additional participants in the AP-1 DNA binding complex in differentiating keratinocytes. AP-1 and CRE DNA binding activity correlated with the induction of CREB, CREMalpha and ATF-1 and CREB phosphorylation at ser133 (ser133 phospho-CREB) in the transition from basal to differentiating keratinocytes, but the activity of a CRE reporter remained unchanged. In contrast, the CRE reporter was activated in the presence of the dominant-negative (DN) CREB mutants, KCREB and A-CREB, proteins that dimerize with CREB family members and block their ability to bind to DNA. The increase in CRE reporter activity in the presence of these mutants suggests that CRE-mediated transcriptional activity is suppressed in keratinocytes through protein-protein interactions involving a factor that dimerizes with the CREB leucine zipper. In experiments where the A-CREB mutant was co-transfected with an AP-1 reporter construct, transcriptional activity was also increased indicating that a CREB family member binds AP-1 sites and represses AP-1 transcriptional activity as well. Exogenous expression of the transcriptional repressor CREMalpha down-regulated both CRE and AP-1 reporters in keratinocytes suggesting that this factor may contribute to the suppression of AP-1 transcriptional activity observed in differentiating keratinocytes.

Animals↗

PubMed1999

Role of CCAAT enhancer-binding protein beta in the thyroid hormone and cAMP induction of phosphoenolpyruvate carboxykinase gene transcription.

Transcription of the gene for phosphoenolpyruvate carboxykinase (PEPCK) is stimulated by thyroid hormone (T3) and cAMP. Two DNA elements in the PEPCK promoter are required for T3 responsiveness including a thyroid hormone response element and a binding site called P3(I) for the CCAAT enhancer-binding protein (C/EBP). Both the alpha and beta isoforms of C/EBP are highly expressed in the liver. C/EBPalpha contributes to the liver-specific expression and cAMP responsiveness of the PEPCK gene. In this study, we examined the ability of C/EBPbeta when bound to the P3(I) site to regulate PEPCK gene expression. We report that C/EBPbeta can stimulate basal expression and participate in the induction of PEPCK gene transcription by T3 and cAMP. The cAMP-responsive element-binding protein and AP1 proteins that contribute to the induction by cAMP are not involved in the stimulation by T3. A small region of the transactivation domain of C/EBPbeta is sufficient for the stimulation of basal expression and cAMP responsiveness. Our results suggest that C/EBPalpha and C/EBPbeta are functionally interchangeable when bound to the P3(I) site of the PEPCK promoter.

Base Sequence↗

PubMed1999

C/EBP regulates the promoter of the eosinophil-derived neurotoxin/RNS2 gene in human eosinophilic cells.

The eosinophil-derived neurotoxin (EDN), a member of the mammalian ribonuclease family, is found in the large specific granules of human eosinophilic leukocytes. We have investigated the role of the C/EBP transcription factor family in the regulation of EDN promoter activity. Here we show that the C/EBP family is involved in intrinsic regulation of EDN promoter activity. We have identified a C/EBP binding site located at -124 in the proximal promoter of the EDN gene. Mutation of this C/EBP site results in a decrease of promoter activity in HL-60-eos cells as well as in eosinophils differentiated in vitro from CD34+ cells. Different C/EBP proteins are able to bind to the C/EBP site as shown by gel shift assay. Our results indicate the importance of the C/EBP family in the regulation of the EDN gene in eosinophils.

Animals↗

PubMed1999

Cell-type-dependent activity of the ubiquitous transcription factor USF in cellular proliferation and transcriptional activation.

USF1 and USF2 are basic helix-loop-helix transcription factors implicated in the control of cellular proliferation. In HeLa cells, the USF proteins are transcriptionally active and their overexpression causes marked growth inhibition. In contrast, USF overexpression had essentially no effect on the proliferation of the Saos-2 osteosarcoma cell line. USF1 and USF2 also lacked transcriptional activity in Saos-2 cells when assayed by transient cotransfection with USF-dependent reporter genes. Yet, there was no difference in the expression, subcellular localization, or DNA-binding activity of the USF proteins in HeLa and Saos-2 cells. Furthermore, Gal4-USF1 and Gal4-USF2 fusion proteins activated transcription similarly in both cell lines. Mutational analysis and domain swapping experiments revealed that the small, highly conserved USF-specific region (USR) was responsible for the inactivity of USF in Saos-2 cells. In HeLa, the USR serves a dual function. It acts as an autonomous transcriptional activation domain at promoters containing an initiator element and also induces a conformational change that is required for USF activity at promoters lacking an initiator. Taken together, these results suggest a model in which the transcriptional activity of the USF proteins, and consequently their antiproliferative activity, is tightly controlled by interaction with a specialized coactivator that recognizes the conserved USR domain and, in contrast to USF, is not ubiquitous. The activity of USF is therefore context dependent, and evidence for USF DNA-binding activity in particular cells is insufficient to indicate USF function in transcriptional activation and growth control.

Amino Acid Sequence↗

PubMed1999

Cyclic adenosine-3',5'-monophosphate-mediated activation of a glutamine synthetase composite glucocorticoid response element.

The glutamate synthetase gene (GS) contains a composite glucocorticoid response element (cGRE) comprised of a GRE and an adjacent element with features of both a cAMP-response element (CRE) and a 12-O-tetradecanoylphorbol 13-acetate (TPA) response element (TRE). The CRE/TRE element of the cGRE contributed to two modes of transcriptional activation: 1) enhancement of the response to cortisol and 2) a synergistic response to cortisol and increased cAMP. COS-7 cells transfected with a cGRE-luciferase construct show minimal expression under basal conditions or forskolin treatment. After cortisol treatment, luciferase activity from the cGRE is enhanced 4- to 8-fold greater than the GRE portion of the cGRE or a GRE from the tyrosine aminotransferase gene. Treatment with both forskolin and cortisol produced a 2- to 4-fold synergistic response over cortisol alone. Synergy is also seen with 8-bromo-cAMP, is specific for the cGRE, and occurs in a number of established cell lines. Elimination of the GRE or CRE/TRE reduces the synergy by 70-100%. Altering the CRE/TRE to GRE spacing changed both enhancement and synergy. Moving the elements 3 bp closer or extending 15 bp reduced enhancement. Synergy was markedly reduced when elements were one half of a helical turn out of phase. Western blots verified that CREB (cAMP-responsive binding protein) and ATF-1 (activating transcription factor-1) binds to the cGRE sequence. A specific dominant negative inhibitor of the CREB family, A-CREB, reduced synergy by 50%. These results suggest that the GS cGRE can potentially integrate signaling from both the cAMP and glucocorticoid receptor transduction pathways and that CREB/ATF-1 may play an important role in this process.

Activating Transcription Factor 1↗

PubMed1998

Life without white fat: a transgenic mouse.

We have generated a transgenic mouse with no white fat tissue throughout life. These mice express a dominant-negative protein, termed A-ZIP/F, under the control of the adipose-specific aP2 enhancer/promoter. This protein prevents the DNA binding of B-ZIP transcription factors of both the C/EBP and Jun families. The transgenic mice (named A-ZIP/F-1) have no white adipose tissue and dramatically reduced amounts of brown adipose tissue, which is inactive. They are initially growth delayed, but by week 12, surpass their littermates in weight. The mice eat, drink, and urinate copiously, have decreased fecundity, premature death, and frequently die after anesthesia. The physiological consequences of having no white fat tissue are profound. The liver is engorged with lipid, and the internal organs are enlarged. The mice are diabetic, with reduced leptin (20-fold) and elevated serum glucose (3-fold), insulin (50- to 400-fold), free fatty acids (2-fold), and triglycerides (3- to 5-fold). The A-ZIP/F-1 phenotype suggests a mouse model for the human disease lipoatrophic diabetes (Seip-Berardinelli syndrome), indicating that the lack of fat can cause diabetes. The myriad of consequences of having no fat throughout development can be addressed with this model.

Adipose Tissue↗

PubMed1998

Inter-helical interactions in the leucine zipper coiled coil dimer: pH and salt dependence of coupling energy between charged amino acids.

We have investigated the physical nature of the observed coupling energy (Delta Delta DeltaGint) between the charged side-chains of the three inter-helical g<-->e' (i, i'+5) pairs (E<-->R, E<-->K, and E<-->E) in the leucine zipper coiled coil dimer. Circular dichroism (CD) spectroscopy measured the thermal stability of eight proteins derived from the basic region leucine zipper domain of chicken VBP, the mammalian TEF at seven pHs and three KCl concentrations. Data from these proteins were used to construct double mutant alanine thermodynamic cycles and determine coupling energies (Delta Delta DeltaGint) for the three g<-->e' pairs. The attractive E<-->R coupling energy of -0.6 kcal mol-1 at low salt decreases to -0.2 kcal mol-1 at high salt. The E<-->K coupling energy of -0.5 kcal mol-1 at low salt decreases to -0.1 kcal mol-1 at high salt. The repulsive E<-->E coupling energy of +0.8 kcal mol-1 at low salt drops to +0.4 at high salt. Reducing the pH to 2.2 halved the attractive coupling energy for the E<-->R and E<-->K pairs while abolishing the repulsion of the E<-->E pair. 13C NMR of a protein selectively labeled with [13Cdelta]glutamate that contained three E<-->R and one R<-->E pair identified four glutamates shifted upfield. We suggest that this is due to electronic perturbation of glutamates in inter-helical E<-->R interactions. Taken together, these data indicate that the E<-->R coupling energy of -0.5 kcal mol-1 at pH 7.4 and 150 mM KCl has an electrostatic component.

Amino Acid Sequence↗

PubMed1998

Coordinate transactivation of the interleukin-2 CD28 response element by c-Rel and ATF-1/CREB2.

The interleukin-2 CD28 response element (CD28RE) acts as a composite enhancer, in conjunction with a 3'-12-O-tetradecanoylphorbol-13-acetate response element (TRE)-like element, to confer CD28 receptor-dependent inducibility to the interleukin-2 promoter in T-cells. When inserted as a single copy upstream of a basal promoter, this composite enhancer, termed the CD28RE-TRE, is both highly active and CD28-inducible in transactivation assays. A multicomponent nuclear protein complex that binds the CD28RE-TRE was isolated by DNA affinity chromatography from nuclear extracts of mitogen- and CD28 receptor-costimulated human T-cells. Immunological and biochemical analyses of this complex reveal the presence of c-Rel, ATF-1, and CREB2 as major DNA-binding components. Coexpression of c-Rel in combination with ATF-1, CREB2, or ATF-1/CREB2 leads to synergistic transactivation of a CD28RE-TRE reporter plasmid in quiescent Jurkat T-cells. Furthermore, CD28-dependent transactivation of the CD28RE-TRE is specifically inhibited by cAMP response element-binding protein (CREB) dominant-negative expression vectors. Moreover, mutant promoter constructs in which the internal 5'-CD28RE and 3'-TRE-like sequences have been topologically positioned 180 degrees out of phase with one another show loss of mitogen- and CD28-dependent inducibility. Finally, the addition of the CREB-binding transcriptional coactivator p300 leads to a dramatic CREB-dependent increase in both mitogen- and CD28-mediated transactivation of the CD28RE-TRE. These findings demonstrate that full physiological responsiveness to CD28 receptor stimulation in T-cells is dependent on topologically linked sequences within the CD28RE-TRE composite enhancer and provide strong support of a direct role for the CREB family of transcription factors and p300/CREB-binding protein coactivator proteins in cytokine gene induction during T-cell activation.

Activating Transcription Factor 1↗

PubMed1998

A dominant-negative inhibitor of CREB reveals that it is a general mediator of stimulus-dependent transcription of c-fos.

Several studies have characterized the upstream regulatory region of c-fos, and identified cis-acting elements termed the cyclic AMP (cAMP) response elements (CREs) that are critical for c-fos transcription in response to a variety of extracellular stimuli. Although several transcription factors can bind to CREs in vitro, the identity of the transcription factor(s) that activates the c-fos promoter via the CRE in vivo remains unclear. To help identify the trans-acting factors that regulate stimulus-dependent transcription of c-fos via the CREs, dominant-negative (D-N) inhibitor proteins that function by preventing DNA binding of B-ZIP proteins in a dimerization domain-dependent fashion were developed. A D-N inhibitor of CREB, termed A-CREB, was constructed by fusing a designed acidic amphipathic extension onto the N terminus of the CREB leucine zipper domain. The acidic extension of A-CREB interacts with the basic region of CREB forming a coiled-coil extension of the leucine zipper and thus prevents the basic region of wild-type CREB from binding to DNA. Other D-N inhibitors generated in a similar manner with the dimerization domains of Fos, Jun, C/EBP, ATF-2, or VBP did not block CREB DNA binding activity, nor did they inhibit transcriptional activation of a minimal promoter containing a single CRE in PC12 cells. A-CREB inhibited activation of CRE-mediated transcription evoked by three distinct stimuli: forskolin, which increases intracellular cAMP; membrane depolarization, which promotes Ca2+ influx; and nerve growth factor (NGF). A-CREB completely inhibited cAMP-mediated, but only partially inhibited Ca2+- and NGF-mediated, transcription of a reporter gene containing 750 bp of the native c-fos promoter. Moreover, glutamate induction of c-fos expression in primary cortical neurons was dependent on CREB. In contrast, induction of c-fos transcription by UV light was not inhibited by A-CREB. Lastly, A-CREB attenuated NGF induction of morphological differentiation in PC12 cells. These results suggest that CREB or its closely related family members are general mediators of stimulus-dependent transcription of c-fos and are required for at least some of the long-term actions of NGF.

Amino Acid Sequence↗

PubMed1998

Green fluorescent protein labeling of cytoskeletal structures--novel targeting approach based on leucine zippers.

Green fluorescent protein (GFP) is a valuable marker for intracellular protein localization. However the fusion of GFP with structural proteins can alter their properties, resulting in a loss of fusion protein localization, decreased GFP fluorescence or both. We describe a novel targeting approach based on noncovalent heterodimerization of GFP and cytoplasmic structural proteins. The formation of structural protein/GFP complexes was mediated by modified leucine zipper protein spacers designed to form high-affinity heterodimers. The complexes localized accurately to specific sites within cells, providing selective fluorescence labeling of subcellular structures such as microfilaments or focal contacts.

Actins↗

PubMed1997

A general method to design dominant negatives to B-HLHZip proteins that abolish DNA binding.

We describe a method to design dominant-negative proteins (D-N) to the basic helix-loop-helix-leucine zipper (B-HLHZip) family of sequence-specific DNA binding transcription factors. The D-Ns specifically heterodimerize with the B-HLHZip dimerization domain of the transcription factors and abolish DNA binding in an equimolar competition. Thermal denaturation studies indicate that a heterodimer between a Myc B-HLHZip domain and a D-N consisting of a 12-amino acid sequence appended onto the Max dimerization domain (A-Max) is -6.3 kcal.mol-1 more stable than the Myc:Max heterodimer. One molar equivalent of A-Max can totally abolish the DNA binding activity of a Myc:Max heterodimer. This acidic extension also has been appended onto the dimerization domain of the B-HLHZip protein Mitf, a member of the transcription factor enhancer binding subfamily, to produce A-Mitf. The heterodimer between A-Mitf and the B-HLHZip domain of Mitf is -3.7 kcal.mol-1 more stable than the Mitf homodimer. Cell culture studies show that A-Mitf can inhibit Mitf-dependent transactivation both in acidic extension and in a dimerization-dependent manner. A-Max can inhibit Myc-dependent foci formation twice as well as the Max dimerization domain (HLHZip). This strategy of producing D-Ns may be applicable to other B-HLHZip or B-HLH proteins because it provides a method to inhibit the DNA binding of these transcription factors in a dimerization-specific manner.

Amino Acid Sequence↗

PubMed1997

Leucine is the most stabilizing aliphatic amino acid in the d position of a dimeric leucine zipper coiled coil.

The energetic contribution of seven amino acids in the d position of a dimeric leucine zipper coiled coil structure was measured by determining the thermal stability. The d position contains the conserved leucines found in the leucine zipper. We used a natural bZIP protein as our host-guest system that remains dimeric when a single d position is mutated. We have determined the thermal stability, monitored by circular dichroism, of 14 proteins which indicate that alanine is 4.6 kcal mol-1 per residue less stabilizing than leucine. The similarly sized amino acid isoleucine is 2.9 kcal mol-1 per residue less stabilizing than leucine, suggesting that leucine is well-packed. Model building indicates that the beta-branched amino acids isoleucine and valine in the d position produced interhelical clashes between the Cgamma2 methyl groups when placed in the favored rotamer conformation. The stabilization by leucine in different d positions is context-dependent; it varies by over 2 kcal mol-1 in the two positions examined. The order of stabilization is L, M, I, V, C, A, and S. Cysteine in the d position can form a disulfide bond which stabilizes the coiled coil.

Amino Acid Sequence↗

PubMed1997

A dominant negative to activation protein-1 (AP1) that abolishes DNA binding and inhibits oncogenesis.

We describe a dominant negative (DN) to activation protein-1 (AP1) that inhibits DNA binding in an equimolar competition. AP1 is a heterodimer of the oncogenes Fos and Jun, members of the bZIP family of transcription factors. The DN, termed A-Fos, consists of a newly designed acidic amphipathic protein sequence appended onto the N-terminus of the Fos leucine zipper, replacing the normal basic region critical for DNA binding. The acidic extension and the Jun basic region form a heterodimeric coiled coil structure that stabilizes the complex over 3000-fold and prevents the basic region of Jun from binding to DNA. Gel shift assays indicate that A-Fos can inactivate the DNA binding of a Fos:Jun heterodimer in an equimolar competition. Transient transfection assays indicate that A-Fos inhibits Jun-dependent transactivation. Both the acidic extension and the Fos leucine zipper are critical for this inhibition. Expression of A-Fos in mouse fibroblasts inhibits focus formation more than colony formation, reflecting the ability of A-Fos to interfere with the AP1 biological functions in mammalian cells. This reagent is more potent than a deletion of either the Fos or Jun transactivation domain, which has been used previously as a dominant negative to AP1 activity.

Amino Acid Sequence↗

PubMed1997

Design of a leucine zipper coiled coil stabilized 1.4 kcal mol-1 by phosphorylation of a serine in the e position.

Using a dimeric bZIP protein, we have designed a leucine zipper that becomes more stable after a serine in the e position is phosphorylated by protein kinase A (delta delta GP = -1.4 kcal mol-1 dimer-1 or -0.7 kcal mol-1 residue-1). Mutagenesis studies indicate that three arginines form a network of inter-helical (i,i' + 5; i, i' + 2) and intra-helical (i, i + 4) attractive interactions with the phosphorylated serine. When the arginines are replaced with lysines, the stabilizing effect of serine phosphorylation is reduced (delta delta GP = -0.5 kcal mol-1 dimer-1). The hydrophobic interface of the leucine zipper needs a glycine in the d position to obtain an increase in stability after phosphorylation. The phosphorylated protein binds DNA with a 15-fold higher affinity. Using a transient transfection assay, we document a PKA dependent four-fold activation of a reporter gene. Phosphorylation of a threonine in the same e position decreases the stability by delta delta GP = +1.2 kcal mol-1 dimer-1. We present circular dichroism (CD) thermal denaturations of 15 bZIP proteins before and after phosphorylation. These data provide insights into the structural determinants that result in stabilization of a coiled coil by phosphorylation.

Amino Acid Sequence↗