Dual function for Tango1 in secretion of bulky cargo and in ER-Golgi morphology.

Tango1 enables ER-to-Golgi trafficking of large proteins. We show here that loss of Tango1, in addition to disrupting protein secretion and ER/Golgi morphology, causes ER stress and defects in cell shape. We find that the previously observed dependence of smaller cargos on Tango1 is a secondary effect. If large cargos like Dumpy, which we identify as a Tango1 cargo, are removed from the cell, nonbulky proteins reenter the secretory pathway. Removal of blocking cargo also restores cell morphology and attenuates the ER-stress response. Thus, failures in the secretion of nonbulky proteins, ER stress, and defective cell morphology are secondary consequences of bulky cargo retention. By contrast, ER/Golgi defects in Tango1-depleted cells persist in the absence of bulky cargo, showing that they are due to a secretion-independent function of Tango1. Therefore, maintenance of ER/Golgi architecture and bulky cargo transport are the primary functions for Tango1.

Revisiting the hydration structure of aqueous Na.

A combination of theory, X-ray diffraction (XRD) and extended x-ray absorption fine structure (EXAFS) are used to probe the hydration structure of aqueous Na+. The high spatial resolution of the XRD measurements corresponds to Qmax = 24 Å-1 while the first-reported Na K-edge EXAFS measurements have a spatial resolution corresponding to 2k = Qmax = 16 Å-1. Both provide an accurate measure of the shape and position of the first peak in the Na-O pair distribution function, gNaO(r). The measured Na-O distances of 2.384 ± 0.003 Å (XRD) and 2.37 ± 0.024 Å (EXAFS) are in excellent agreement. These measurements show a much shorter Na-O distance than generally reported in the experimental literature (Na-Oavg ∼ 2.44 Å) although the current measurements are in agreement with recent neutron diffraction measurements. The measured Na-O coordination number from XRD is 5.5 ± 0.3. The measured structure is compared with both classical and first-principles density functional theory (DFT) simulations. Both of the DFT-based methods, revPBE and BLYP, predict a Na-O distance that is too long by about 0.05 Å with respect to the experimental data (EXAFS and XRD). The inclusion of dispersion interactions (-D3 and -D2) significantly worsens the agreement with experiment by further increasing the Na-O distance by 0.07 Å. In contrast, the use of a classical Na-O Lennard-Jones potential with SPC/E water accurately predicts the Na-O distance as 2.39 Å although the Na-O peak is over-structured with respect to experiment.

Dressed adiabatic and diabatic potentials to study topological effects for F + H2.

We report here on the two lowest, rigorous-accurate diabatic potential energy surfaces (PES), for the F + H2 system, as calculated by including the two dominant topological effects of this system at the low energy region, namely, the Jahn-Teller effect and the Renner-Teller effect. Both effects were treated in the most rigorous way as demanded by the Born-Oppenheimer approach. No approximations were made, and in those cases where convergence was required, it was satisfied. In other words, convergence was attained in all situations. The numerical part that includes the calculation of the two lowest ab initio adiabatic PESs and the corresponding nonadiabatic coupling terms (NACTs) was carried out using the MOLPRO program. The required diabatic potentials are calculated by employing these ab initio adiabatic PESs and the corresponding adiabatic-to-diabatic angles as obtained employing the above-mentioned ab initio NACTs. The relevance of these Renner-Teller/Jahn-Teller diabatic potentials is studied by comparing the dressed-lowest ab initio adiabatic PES and the one formed by diagonalizing the dressed-diabatic 2 × 2 potential matrix. The dressed-potentials are calculated employing the vib-rotational manifold derived for each of the three surfaces, namely, the lowest adiabatic potential and the two diabatic ones. This kind of study was recently recommended by Lipoff and Herschbach ( Mol. Phys. 2010 , 108 , 1133 ) as a "blessed-practice" for the relevance of any PES. In the present case significant differences were revealed between the two types of dressed-adiabatic PESs, eventually, indicating that the lowest, ab initio PES (due to the Born-Oppenheimer approximation) is not adequate for low energy processes.

A tri-atomic Renner-Teller system entangled with Jahn-Teller conical intersections.

The present study concentrates on a situation where a Renner-Teller (RT) system is entangled with Jahn-Teller (JT) conical intersections. Studies of this type were performed in the past for contours that surround the RT seam located along the collinear axis [see, for instance, G. J. Halász, Á. Vibók, R. Baer, and M. Baer, J. Chem. Phys. 125, 094102 (2006)]. The present study is characterized by planar contours that intersect the collinear axis, thus, forming a unique type of RT-non-adiabatic coupling terms (NACT) expressed in terms of Dirac-δ functions. Consequently, to calculate the required adiabatic-to-diabatic (mixing) angles, a new approach is developed. During this study we revealed the existence of a novel molecular parameter, η, which yields the coupling between the RT and the JT NACTs. This parameter was found to be a pure number η = 22/π (and therefore independent of any particular molecular system) and is designated as Renner-Jahn coupling parameter. The present study also reveals an unexpected result of the following kind: It is well known that each (complete) group of states, responsible for either the JT-effect or the RT-effect, forms a Hilbert space of its own. However, the entanglement between these two effects forms a third effect, namely, the RT/JT effect and the states that take part in it form a different Hilbert space.

Symptomatic response to imatinib mesylate in cutaneous mastocytosis associated with chronic myelomonocytic leukemia.

Mastocytosis is an uncommon disorder defined by increased and abnormal mast cells in one or more tissues. Cutaneous mastocytosis (cm) is limited to the skin, with varying degrees of rash, pruritus, and disfigurement. Systemic mastocytosis (sm) typically involves the bone marrow, sometimes in association with other bone marrow disorders, including chronic myelomonocytic leukemia (cmml). Mastocytosis has been associated with somatic mutations in the gene encoding the tyrosine kinase Kit, leading to identification of Kit as a therapeutic target. The Kit inhibitor imatinib mesylate is approved for aggressive sm. We present an unusual patient with disabling pruritus from telangiectasia macularis eruptiva perstans, a subtype of cm, and cmml, but with no evidence of systemic mast cell disease. She was treated with imatinib and experienced marked improvement in her pruritus. Concomitant cm and cmml have not previously been reported, and the present report is the first of successful imatinib therapy in an adult patient with cm.

PcrV antibody-antibiotic combination improves survival in Pseudomonas aeruginosa-infected mice.

The type III secretion system (TTSS) of Pseudomonas aeruginosa, associated with acute infection, facilitates the direct injection of cytotoxins into the host cell cytoplasm. Mab166, a murine monoclonal antibody against PcrV, a protein located at the tip of the injectisome, has demonstrated efficacy against P. aeruginosa infection, resulting in reduced lung injury and increased survival in murine models of infection. We hypothesised that the administration of Mab166 in combination with an antibiotic would further improve the survival of P. aeruginosa-infected mice. A murine model of P. aeruginosa acute infection, three clinically relevant antibiotics (ciprofloxacin, tobramycin and ceftazidime) and the Mab166 antibody were used for this study. Consistently, compared to other treatment groups (antibiotic or antibody administered in isolation), the combination of Mab166 and antibiotic significantly improved the survival of mice infected with three times the lethal dose (LD(90)) of the highly cytotoxic ExoU-secreting strain, PA103. This synergistic effect was primarily due to enhanced bactericidal effect and protection against lung injury, which prevented bacterial dissemination to other organs. Hence, the combination of Mab166 with antibiotic administration provides a new, more effective strategy against P. aeruginosa airway infection, especially when large numbers of highly virulent strains are present.

Longitudinal alterations in mRNA expression of the BDNF neurotrophin signaling cascade in blood correlate with changes in depression scores in patients undergoing electroconvulsive therapy.

Electroconvulsive therapy (ECT) appears to be the most effective treatment for severe depression. However, its mechanisms of action are incompletely understood. Evidence suggests ECT enhances neuroplasticity and neurogenesis. While studies on ECT-induced neuroplasticity focused on brain-derived neurotrophic factor (BDNF), other factors of the BDNF/TrkB signaling cascade remain underinvestigated. We assessed longitudinal changes in depression scores, serum BDNF protein levels, and mRNA expression of BDNF/TrkB related genes (BDNF, AKT1, ERK1, CREB), NR3C1 and IGF1 in peripheral blood in 19 treatment-resistant depressed patients undergoing ECT. We also analysed DNA methylation patterns at various timepoints to explore possible epigenetic regulation of mRNA expression. Using multilevel regression, we found a negative association between depression scores and blood-based mRNA expression of BDNF/TrkB related genes and NR3C1. Expression of BDNF, ERK1 and NR3C1 increased significantly over time (BDNF: ß = 0.0295, p = 0.003; ERK1: ß = 0.0170, p = 0.034; NR3C1: ß = 0.0035, p = 0.050). For these three genes changes in mRNA expression were highly correlated (R = 0.59 - 0.88) with changes in DNA methylation for multiple CpG sites in the respective genes. Also, serum BDNF protein levels increased across the study period (ß = 0.11, p = 0.001). Our findings show that the antidepressant effects of ECT are associated with changes in expression of BDNF and its signaling molecules and that these molecular markers can be detected in peripheral blood. Alterations in DNA methylation could be a key mechanism whereby ECT influences gene expression.

Granulomatous amebic encephalitis: an under-recognized cause of infectious mortality after hematopoietic stem cell transplantation.

Granulomatous amebic encephalitis (GAE) is a rare, nearly always fatal form of encephalitis that occurs mostly in the setting of immune compromise or chronic disease. The prevalence and clinical characteristics of this Acanthamoeba infection in hematopoietic stem cell transplant (HSCT) recipients are not well described. We present an HSCT patient in whom the diagnosis of GAE was made at autopsy. A systematic review of previously reported cases is provided to highlight the clinical presentation and early diagnostic features of GAE in HSCT recipients. Amebic infection usually initially involves the skin or lungs over a period of months, and becomes rapidly fatal once it crosses the blood-brain barrier. GAE is usually discovered postmortem owing to lack of awareness of this deadly infection and delay in diagnosis. Subacute presentation of multiple recurrent panniculitis-like subcutaneous nodules associated with eosinophilia and a history of chronic rhinitis or sinusitis warrant investigation for a possible amebic infection. Prolonged corticosteroid use and a recent exposure to unhygienic water are potential risk factors for GAE. Successful outcomes may be achieved with early intensive treatment using a combination of effective drugs.

Successful electroconvulsive therapy for depression in a man with cerebral amyloid angiopathy.

Cerebral amyloid angiopathy (CAA) is a condition characterised by accumulation of amyloid beta protein (Aß) in the wall of cerebral blood vessels which increases the risk of intracranial haemorrhage and contributes to cognitive impairment. We describe the case of a man around the age of 70 with 'probable' CAA according to the modified Boston criteria and severe depression whose depression was treated successfully with electroconvulsive therapy (ECT). To the best of our knowledge, there are no earlier published reports of ECT in a patient with CAA. We briefly discuss possible safety measures for these patients, the impact of ECT on cognition in CAA and a possible influence of ECT on Aß clearance.

Molecular symmetry properties of conical intersections and nonadiabatic coupling terms: theory and quantum chemical demonstration for cyclopenta-2,4-dienimine (C5H4NH).

This paper discovers molecular symmetry (MS) properties of conical intersections (CIs) and the related nonadiabatic coupling terms (NACTs) in molecules which allow large amplitude motions such as torsion, in the frame of the relevant molecular symmetry group, focusing on groups with one-dimensional (1-d) irreducible representations (IREPs). If one employs corresponding MS-adapted nuclear coordinates, the NACTs can be classified according to those IREPs. The assignment is supported by theorems which relate the IREPs of different NACTs to each other, and by properties of the NACTs related to the CIs. For example, planar contour integrals of the NACTs evaluated along loops around the individual CIs are equal to +pi or -pi, depending on the IREP-adapted signs of the NACTs. The + or - signs for the contour integrals may also be used to define the "charges" and IREPs of the CIs. We derive various general molecular symmetry properties of the related NACTs and CIs. These provide useful applications; e.g., the discovery of an individual CI allows one to generate, by means of all molecular symmetry operations, the complete set of CIs at different symmetry-related locations. Also, we show that the seams of CIs with different IREPs may have different topologies in a specific plane of MS-adapted coordinates. Moreover, the IREPs impose symmetrical nodes of the NACTs, and this may support their calculations by quantum chemical ab initio methods, even far away from the CIs. The general approach is demonstrated by application to an example. Specifically, we investigate the CIs and NACTs of cyclopenta-2,4-dienimine (C(5)H(4)NH) which has C(2v)(M) molecular symmetry with 1-d IREPs. The results are confirmed by quantum chemical calculations, starting from the location of a CI based on the Longuet-Higgins phase change theorem, until a proof of self-consistency, i.e., the related symmetry-adapted NACTs fulfill quantization rules which have been derived in [Baer, M. Beyond Born-Oppenheimer: Electronic non-Adiabatic Coupling Terms and Conical Intersections; Wiley & Sons Inc.: Hoboken, NJ, 2006].

A catalytic RNA and its gene from Salmonella typhimurium.

The gene for the RNA subunit (M1 RNA) of ribonuclease P from Salmonella typhimurium directs the synthesis of an RNA that can cleave transfer RNA precursor molecules. The mature M1 RNA coded for by Salmonella typhimurium is 375 nucleotides long and has six nucleotide changes in comparison to M1 RNA from Escherichia coli. The regions for promotion and termination of transcription are closely conserved, but adjacent regions of nucleotide sequences show considerable drift.

Scavenger receptor A is expressed by macrophages in response to Porphyromonas gingivalis, and participates in TNF-alpha expression.

INTRODUCTION: Porphyromonas gingivalis is a periodontopathic bacterium closely associated with generalized aggressive periodontal disease. Pattern recognition receptors (PRRs) participate in host response to this organism. It is likely that PRRs not previously recognized as part of the host response to P. gingivalis also participate in host response to this organism. METHODS AND RESULTS: Employing qRT-PCR, we observed increased msr1 gene expression at 2, 6, and 24 h of culture with P. gingivalis strain 381. Flow cytometry revealed increased surface expression of SR-A protein by the 24 h time point. Macrophages cultured with an attachment impaired P. gingivalis fimA- mutant (DPG3) expressed intermediate levels of SR-A expression. Heat-killed P. gingivalis stimulated SR-A expression similar to live bacteria, and purified P. gingivalis capsular polysaccharide stimulated macrophage SR-A expression, indicating that live whole organisms are not necessary for SR-A protein expression in macrophage response. As SR-A is known to play a role in lipid uptake by macrophages, we tested the ability of low-density lipoprotein (LDL) to influence the SR-A response of macrophages to P. gingivalis, and observed no effect of LDL on P. gingivalis-elicited SR-A expression. Lastly, we observed that SR-A knockout (SR-A(-/-)) mouse macrophages produced significantly more tumor necrosis factor (TNF)-alpha than wild type mouse macrophages cultured with P. gingivalis. CONCLUSION: These data identify that SR-A is expressed by macrophages in response to P. gingivalis, and support that this molecule plays a role in TNF-alpha production by macrophages to this organism.

Intralines of quasi-conical intersections on torsion planes: methylamine as a case study.

Recently we reported on a novel feature associated with the intersection of the two lowest states (1)A' and (1)A'' of the methylamine (J. Chem. Phys. 2008, 128, 244302). We established the existence of a finite (closed) line of conical intersections (ci), namely, a finite seam, located in the HC-NHH symmetry plane, a line that is formed by moving a single hydrogen on that plane while locking the positions of the (six) other atoms. In the present article, this study is extended to the corresponding torsion planes formed by rotating the methyl group around the CN axis. The torsion planes, in contrast with the HC-NHH symmetry plane, do not satisfy the symmetry feature that enables the seam just mentioned. Nevertheless, the calculated nonadiabatic coupling terms (NACTs) resemble features similar to those encountered in the HC-NHH symmetry plane. Following a tedious numerical study supported by a theoretical model (Section III), it was verified that these NACTs may become similar to those on the symmetry plane, sometimes even to the level of almost no distinction, but lack one basic feature; namely, they are not singular and therefore do not form topological effects.

Efficacy of decitabine in the treatment of patients with chronic myelomonocytic leukemia (CMML).

Chronic myelomonocytic leukemia (CMML) characterized by cytopenias, bone marrow and peripheral blood cell dysplasia is notoriously hard to treat. Recent reclassification of CMML as a myelodysplastic/myeloproliferative (MDS/MPS) disease rather than a myelodysplastic syndrome (MDS) by the World Health Organisation (WHO) has led to a review of CMML patients treated with decitabine. Overall response rates (ORR) (complete response [CR]+partial response [PR]) in the subset of patients with CMML in one pivotal phase 3 trial (D-0007) and two phase 2 trials (PCH 95-11, PCH 97-19) decitabine were reviewed. For consistency across trials, all decitabine-treated patients were evaluated using the phase 2 response criteria (CR was defined by normocellular bone marrow with <5% blasts and normal Hgb, WBC, and platelet counts, and PR required 50% decrease in blast count, increases in Hgb by >1.5 mmol/L, WBC count by >1000, and platelet count by >50,000). A total of 31 patients diagnosed with CMML are included in this review. Similar demographics and disease characteristics were observed in all three studies, with an average age of 70.2 years and 71% of patients male. Baseline WBC of >20,000 were observed in 8/28 (29%) patients and baseline bone marrow blasts >5% in 11/28 (39%) patients. All clinical responses were centrally reviewed. The ORR was 25% (14% CR+11% PR). Hematologic improvement was observed in 11% of patients and stable disease in 39% of patients. The decitabine adverse event profile seen in CMML patients was similar to observations in other hematologic patient populations, with myelosuppression and related infectious complications. These data demonstrate encouraging activity for decitabine in CMML, and suggest that studies in other myeloproliferative diseases may be warranted.

An intraline of conical intersections for methylamine.

In this article are considered the conical intersections (ci's) related to the N-H bond in the methylamine, CH(3)NH(2), molecule. The novel feature that was revealed is that the two lowest states 1A(') and 1A(") are coupled by a line of cis located in HC-NHH plane-a line that is formed by moving a single hydrogen on that plane while fixing the (six) other atoms. The validity of this line was proven first by studying the singularities of the (angular) nonadiabatic coupling terms and then by revealing the degeneracy points formed by the two interacting adiabatic potential energy surfaces (PESs). A theoretical analysis indicated that the line has to be a finite closed line. We also calculated the Berry phase for a contour that surrounds this line and found it to be 3.127 rad, namely, a value reasonably close to pi. The existence of such lines of cis-instead of isolated cis (as exhibited by other n-atomic (n>3) molecules such as HNCO or C(2)H(2))-may enhance significantly the transition rate from an upper adiabatic state to a lower one. There are also numerical advantages in such situations, that is, if such a line is properly placed in that plane (like in the present case) the wave-packet treatment of the nuclei can be carried out employing a single diabatic PES instead of having to consider two coupled PESs.

A compact strip-line pulsed power generator for isentropic compression experiments.

Veloce is a medium-voltage, high-current, compact pulsed power generator developed for isentropic and shock compression experiments. Because of its increased availability and ease of operation, Veloce is well suited for studying isentropic compression experiments (ICE) in much greater detail than previously allowed with larger pulsed power machines such as the Z accelerator. Since the compact pulsed power technology used for dynamic material experiments has not been previously used, it is necessary to examine several key issues to ensure that accurate results are obtained. In the present experiments, issues such as panel and sample preparation, uniformity of loading, and edge effects were extensively examined. In addition, magnetohydrodynamic simulations using the ALEGRA code were performed to interpret the experimental results and to design improved sample/panel configurations. Examples of recent ICE studies on aluminum are presented.

Phase 2 study of gandotinib (LY2784544) in patients with myeloproliferative neoplasms.

BACKGROUND: The Philadelphia chromosome-negative myeloproliferative neoplasms (MPNs) are associated with increases in janus kinase 2 (JAK2) signaling, often resulting from the JAK2 V617F mutation. LY2784544 (gandotinib) is a potent, selective, small-molecule inhibitor of JAK2 that has potential dose-dependent selectivity for the JAK2 V617F mutation and may inhibit additional JAK2 mutant isoforms in nonclinical testing. METHODS: A multicenter, single-arm, outpatient phase 2 study evaluated the efficacy, safety, and pharmacokinetics (PK) of gandotinib administered to patients (120 mg once daily) with MPNs, including polycythemia vera (PV), essential thrombocythemia (ET), and myelofibrosis (MF). Between May 2012 and March 2015, 138 patients received at least one dose of study drug. FINDINGS: Most frequent Grade 3 or 4 treatment-emergent adverse events that were considered study-drug related were anemia (11.6%), hyperuricemia (3.2%), fatigue (2.9%), diarrhea (2.2%), and thrombocytopenia (2.2%). Overall response rates (ORRs) in patients with JAK2 V617F-mutated PV, ET, and MF were 95%, 90.5%, and 9.1%, respectively, while patients with ET and MF without the JAK2 V617F mutations had ORRs of 43.7% and 0%, respectively. INTERPRETATIONS: LY2784544 demonstrated efficacy in JAK2 V617F-mutated MPNs, including in patients previously on ruxolitinib therapy, who had an ORR of 3.3%. At the 1-year visit, 44% of patients experienced a ≥50% improvement in the MPN-Symptom Assessment Form Total Symptom Score, and 26% of patients had a 50% reduction in Brief Fatigue Inventory score.

Human Ste20 homologue hPAK1 links GTPases to the JNK MAP kinase pathway.

BACKGROUND: The Rho-related GTP-binding proteins Cdc42 and Rac1 have been shown to regulate signaling pathways involved in cytoskeletal reorganization and stress-responsive JNK (Jun N-terminal kinase) activation. However, to date, the GTPase targets that mediate these effects have not been identified. PAK defines a growing family of mammalian kinases that are related to yeast Ste20 and are activated in vitro through binding to Cdc42 and Rac1 (PAK: p21 Cdc42-/Rac-activated kinase). Clues to PAK function have come from studies of Ste20, which controls the activity of the yeast mating mitogen-activated protein (MAP) kinase cascade, in response to a heterotrimeric G protein and Cdc42. RESULTS: To initiate studies of mammalian Ste20-related kinases, we identified a novel human PAK isoform, hPAK1. When expressed in yeast, hPAK1 was able to replace Ste20 in the pheromone response pathway. Chemical mutagenesis of a plasmid encoding hPAK1, followed by transformation into yeast, led to the identification of a potent constitutively active hPAK1 with a substitution of a highly conserved amino-acid residue (L107F) in the Cdc42-binding domain. Expression of the hPAK1(L107F) allele in mammalian cells led to specific activation of the Jun N-terminal kinase MAP kinase pathway, but not the mechanistically related extracellular signal-regulated MAP kinase pathway. CONCLUSIONS: These results demonstrate that hPAK1 is a GTPase effector controlling a downstream MAP kinase pathway in mammalian cells, as Ste20 does in yeast. Thus, PAK and Ste20 kinases play key parts in linking extracellular signals from membrane components, such as receptor-associated G proteins and Rho-related GTPases, to nuclear responses, such as transcriptional activation.

Photolyases from Saccharomyces cerevisiae and Escherichia coli recognize common binding determinants in DNA containing pyrimidine dimers.

DNA photolyases catalyze the light-dependent repair of pyrimidine dimers in DNA. The results of nucleotide sequence analysis and spectroscopic studies demonstrated that photolyases from Saccharomyces cerevisiae and Escherichia coli share 37% amino acid sequence homology and contain identical chromophores. Do the similarities between these two enzymes extend to their interactions with DNA containing pyrimidine dimers, or does the organization of DNA into nucleosomes in S. cerevisiae necessitate alternative or additional recognition determinants? To answer this question, we used chemical and enzymatic techniques to identify the contacts made on DNA by S. cerevisiae photolyase when it is bound to a pyrimidine dimer and compared these contacts with those made by E. coli photolyase and by a truncated derivative of the yeast enzyme when bound to the same substrate. We found evidence for a common set of interactions between the photolyases and specific phosphates in the backbones of both strands as well as for interactions with bases in both the major and minor grooves of dimer-containing DNA. Superimposed on this common pattern were significant differences in the contributions of specific contacts to the overall binding energy, in the interactions of the enzymes with groups on the complementary strand, and in the extent to which other DNA-binding proteins were excluded from the region around the dimer. These results provide strong evidence both for a conserved dimer-binding motif and for the evolution of new interactions that permit photolyases to also act as accessory proteins in nucleotide excision repair. The locations of the specific contacts made by the yeast enzyme indicate that the mechanism of nucleotide excision repair in this organism involves incision(s) at a distance from the pyrimidine dimer.

The ability of C/EBP beta but not C/EBP alpha to synergize with an Sp1 protein is specified by the leucine zipper and activation domain.

The rat CYP2D5 P-450 gene is activated in the liver during postnatal development. We previously showed that liver-specific transcription of the CYP2D5 gene is dictated by a proximal promoter element, termed 2D5, that is composed of a binding site for Sp1 or a related factor, and an adjacent cryptic C/EBP (CCAAT/enhancer-binding protein) site. Despite the fact that both C/EBP alpha and C/EBP beta are expressed abundantly in liver, only C/EBP beta is capable of stimulating the 2D5 promoter in HepG2 hepatocarcinoma cells. In addition, activation of the 2D5 promoter by C/EBP beta is completely dependent on the presence of the Sp1 site. Domain switch experiments reveal that C/EBP beta proteins containing either the leucine zipper or the activation domain of C/EBP alpha are unable to stimulate the 2D5 promoter yet are fully capable of transactivating an artificial promoter bearing a high-affinity C/EBP site. Thus, the leucine zipper and the activation domain of C/EBP beta are absolutely required to support transactivation of the 2D5 promoter. Using Drosophila cells that lack endogenous Sp1 activity, we show that the serine/threonine- and glutamine-rich activation domains A and B of Sp1 are required for efficient cooperatively with C/EBP beta. Furthermore, analysis of c/ebp beta-deficient mice shows that mutant animals are defective in expression of a murine CYP2D5 homolog in hepatic cells, confirming the selective ability of C/EBP beta to activate this liver-specific P-450 gene in vivo. Our findings illustrate that two members of a transcription factor family can achieve distinct target gene specificities through differential interactions with a cooperating Sp1 protein.