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The scientific publication landscape is changing quickly, with an enormous increase in options and models. Articles can be published in a complex variety of journals that differ in their presentation format (online-only or in-print), editorial organizations that maintain them (commercial and/or society-based), editorial handling (academic or professional editors), editorial board composition (academic or professional), payment options to cover editorial costs (open access or pay-to-read), indexation, visibility, branding, and other aspects. Additionally, online submissions of non-revised versions of manuscripts prior to seeking publication in a peer-reviewed journal (a practice known as pre-printing) are a growing trend in biological sciences. In this changing landscape, researchers in biochemistry and molecular biology must re-think their priorities in terms of scientific output dissemination. The evaluation processes and institutional funding for scientific publications should also be revised accordingly. This article presents the results of discussions within the Department of Biochemistry, University of São Paulo, on this subject.

Lung cancer is the most common cause of cancer deaths. The expression of the transcription factor C/EBPα (CCAAT/enhancer binding protein α) is frequently lost in non-small cell lung cancer, but the mechanisms by which C/EBPα suppresses tumor formation are not fully understood. In addition, no pharmacological therapy is available to specifically target C/EBPα expression. We discovered a subset of pulmonary adenocarcinoma patients in whom negative/low C/EBPα expression and positive expression of the oncogenic protein BMI1 (B lymphoma Mo-MLV insertion region 1 homolog) have prognostic value. We also generated a lung-specific mouse model of C/EBPα deletion that develops lung adenocarcinomas, which are prevented by Bmi1 haploinsufficiency. BMI1 activity is required for both tumor initiation and maintenance in the C/EBPα-null background, and pharmacological inhibition of BMI1 exhibits antitumor effects in both murine and human adenocarcinoma lines. Overall, we show that C/EBPα is a tumor suppressor in lung cancer and that BMI1 is required for the oncogenic process downstream of C/EBPα loss. Therefore, anti-BMI1 pharmacological inhibition may offer a therapeutic benefit for lung cancer patients with low expression of C/EBPα and high BMI1., (Copyright © 2016, American Association for the Advancement of Science.)

The serine/threonine protein kinase Akt promotes cell survival, growth, and proliferation through phosphorylation of different downstream substrates. A key effector of Akt is the mammalian target of rapamycin (mTOR). Akt is known to stimulate mTORC1 activity through phosphorylation of tuberous sclerosis complex 2 (TSC2) and PRAS40, both negative regulators of mTOR activity. We previously reported that IκB kinase α (IKKα), a component of the kinase complex that leads to NF-κB activation, plays an important role in promoting mTORC1 activity downstream of activated Akt. Here, we demonstrate IKKα-dependent regulation of mTORC1 using multiple PTEN null cancer cell lines and an animal model with deletion of IKKα. Importantly, IKKα is shown to phosphorylate mTOR at serine 1415 in a manner dependent on Akt to promote mTORC1 activity. These results demonstrate that IKKα is an effector of Akt in promoting mTORC1 activity., (© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.)

Purpose: We sought to determine the mechanisms of downregulation of the airway transcription factor Foxa2 in lung cancer and the expression status of Foxa2 in non-small-cell lung cancer (NSCLC)., Methods: A series of 25 lung cancer cell lines were evaluated for Foxa2 protein expression, FOXA2 mRNA levels, FOXA2 mutations, FOXA2 copy number changes and for evidence of FOXA2 promoter hypermethylation. In addition, 32 NSCLCs were sequenced for FOXA2 mutations and 173 primary NSCLC tumors evaluated for Foxa2 expression using an immunohistochemical assay., Results: Out of the 25 cell lines, 13 (52%) had undetectable FOXA2 mRNA. The expression of FOXA2 mRNA and Foxa2 protein were congruent in 19/22 cells (p = 0.001). FOXA2 mutations were not identified in primary NSCLCs and were infrequent in cell lines. Focal or broad chromosomal deletions involving FOXA2 were not present. The promoter region of FOXA2 had evidence of hypermethylation, with an inverse correlation between FOXA2 mRNA expression and presence of CpG dinucleotide methylation (p < 0.0001). In primary NSCLC tumor specimens, there was a high frequency of either absence (42/173, 24.2%) or no/low expression (96/173, 55.4%) of Foxa2. In 130 patients with stage I NSCLC there was a trend towards decreased survival in tumors with no/low expression of Foxa2 (HR of 1.6, 95%CI 0.9-3.1; p = 0.122)., Conclusions: Loss of expression of Foxa2 is frequent in lung cancer cell lines and NSCLCs. The main mechanism of downregulation of Foxa2 is epigenetic silencing through promoter hypermethylation. Further elucidation of the involvement of Foxa2 and other airway transcription factors in the pathogenesis of lung cancer may identify novel therapeutic targets., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

The transcription factor RUNX1 is essential to establish the haematopoietic gene expression programme; however, the mechanism of how it activates transcription of haematopoietic stem cell (HSC) genes is still elusive. Here, we obtained novel insights into RUNX1 function by studying regulation of the human CD34 gene, which is expressed in HSCs. Using transgenic mice carrying human CD34 PAC constructs, we identified a novel downstream regulatory element (DRE), which is bound by RUNX1 and is necessary for human CD34 expression in long-term (LT)-HSCs. Conditional deletion of Runx1 in mice harbouring human CD34 promoter-DRE constructs abrogates human CD34 expression. We demonstrate by chromosome conformation capture assays in LT-HSCs that the DRE physically interacts with the human CD34 promoter. Targeted mutagenesis of RUNX binding sites leads to perturbation of this interaction and decreased human CD34 expression in LT-HSCs. Overall, our in vivo data provide novel evidence about the role of RUNX1 in mediating interactions between distal and proximal elements of the HSC gene CD34.

Ankyrin defects are the most common cause of hereditary spherocytosis (HS). In several kindreds with recessive, ankyrin-deficient HS, mutations have been identified in the ankyrin promoter that have been proposed to decrease ankyrin synthesis. We analyzed the effects of two mutations, -108T to C and -108T to C in cis with -153G to A, on ankyrin expression. No difference between wild type and mutant promoters was demonstrated in transfection or gel shift assays in vitro. Transgenic mice with a wild type ankyrin promoter linked to a human (A)gamma-globin gene expressed gamma-globin in 100% of erythrocytes in a copy number-dependent, position-independent manner. Transgenic mice with the mutant -108 promoter demonstrated variegated gamma-globin expression, but showed copy number-dependent and position-independent expression similar to wild type. Severe effects in ankyrin expression were seen in mice with the linked -108/-153 mutations. Three transgenic lines had undetectable levels of (A)gamma-globin mRNA, indicating position-dependent expression, and four lines expressed significantly lower levels of (A)gamma-globin mRNA than wild type. Two of four expressing lines showed variegated gamma-globin expression, and there was no correlation between transgene copy number and RNA level, indicating copy number-independent expression. These data are the first demonstration of functional defects caused by HS-related, ankyrin gene promoter mutations.

Hereditary spherocytosis (HS) is a common hemolytic anemia caused by defects in the erythrocyte membrane proteins. The screening of mutations in the ankyrin-1 (ANK1) gene of 28 Brazilian HS patients showed two new missense mutations (His276Arg and Ile1054Thr) and one novel promoter mutation (-153 G-->A). The His276Arg mutation affected the invariable TPLH sequence on repeat 9. The -153 mutation was linked in cis to the known -108 T-->C mutation. In contrast to other populations, we were able to detect mutations in the ankyrin-1 gene in only 10% of our patients. It is also interesting to point out that, from 15 informative subjects for the 3' Acn repeats, only one presented a loss of heterozigosity at the cDNA level. Taken together, these results suggest that mutations in the ankyrin-1 gene might not be as common in Brazil as described for other populations., (Copyright 2000 Wiley-Liss, Inc.)

We analyzed the frequency of the C282Y and H63D mutations in the HFE gene in 227 individuals from Brazil comprising 71 Caucasians, 91 racially mixed Caucasian African-derived Amerindians (both populations from Southeast Brazil), 85 African-derived subjects (from Northeast Brazil) and 75 Parakanã Indians. Allelic frequency of the mutation C. 845G(A (C282Y) was 1.4% in the Caucasian population, 1.1% in the African-derived population, 1.1% in the racially mixed normal controls and 0% in the Parakanã Indians. In the African-derived population, the C282Y mutation was present on chromosomes bearing the haplotype 6/1h according to Beutler and West (1997). Allelic frequency of the mutation C. 187C(G (H63D) was 16.3% in the Caucasian population, 7.5% in the African-derived population, 9.8% in the racially mixed controls and 0% in the Amerindians. The presence of these mutations in the African-derived population reflects the fact that these subjects may have undergone a non-identified racial admixture in their past history. The absence of both defects in the Amerindians suggests that these mutations have emerged after the migration of Polynesians to America, or that they may not have reached the Polynesian population until after the migration to America had occurred., (Copyright 1999 Academic Press.)

We report the clinical and laboratory findings in three unrelated families from southeastern Brazil with Sp alpha I/65 hereditary elliptocytosis (HE), including one homozygote and a patient presenting an elongated beta-spectrin. In family 1, three patients presented the allele alpha-Lely in trans to the elliptocytogenic allele. In these three patients the blood smear showed pronounced elliptocytosis, poikilocytosis and a few small red cell fragments instead of the mild elliptocytosis observed in their father, who did not present the polymorphism. In family 2 we describe one homozygote, with consanguineous parents presenting with anaemia, splenomegaly, severe poikilocytosis and elliptocytosis, budding, microspherocytes and numerous fragments in the blood smear. In family 3 we found an elongated beta Sp in a patient with Sp alpha I/65. The cause of the HE was the Sp alpha I/65 since the elongated beta Sp was not found in his brother, who also presented with HE and Sp alpha I/65. Apparently the abnormal beta Sp did not aggravate the HE, because both individuals had the same clinical and laboratory findings. However, the propositus presented a few more elliptocytes and poikilocytes than his brother, probably because the elongated beta-spectrin may have disturbed the spectrin self-association. In fact, in the propositus an abnormal band was observed in the nondenaturing gels, just above the Sp dimer, probably as a result of the association of the abnormal beta Sp with the normal spectrin chains. In the family studied here, both brothers presented the allele alpha Lely, but as their mother was dead, it was not possible to determine the polymorphism transmission. However, the high number of poikilocytes observed in the blood smear of both cases suggests an association in trans with the Sp alpha I/65. Thus, taken together, the data in this report indicate that HE secondary to Sp alpha I/65 abnormality is frequent in Brazil, and in one case it was associated with an apparently novel abnormal large beta-spectrin.

We have recently shown that the permeabilization of the inner mitochondrial membrane by Ca2+ plus prooxidants is associated with oxidation of protein thiols forming cross-linked protein aggregates (Fagian, M.M., Pereira-da-Silva, L., Martins, I.S. and Vercesi, A.E. (1990) J. Biol. Chem. 265, 19955-19960). In this study we show that the incubation of rat liver mitochondria in the presence of the thiol reagent 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) and Ca2+ caused production of membrane protein aggregates, mitochondrial swelling, disruption of membrane potential and Ca2+ release. The presence of DTT prevented but did not reverse the elimination of delta psi induced by DIDS. EGTA prevented delta psi elimination and decreased the amount of protein aggregates, suggesting that the binding of Ca2+ to some membrane protein may expose buried thiols to react with DIDS. Reversal of collapsed delta psi by EGTA indicates that DIDS-induced protein aggregates require the presence of Ca2+ for significant membrane permeabilization. Cyclosporin A prevented mitochondrial swelling, suggesting that DIDS-induced membrane protein polymerization mimics the condition designated as Ca(2+)-induced permeabilization transition of mitochondria. The lack of oxidation of pyridine nucleotides or significant lipid peroxidation by DIDS supports the notion that membrane permeabilization by this compound is mediated by its interaction with membrane proteins.