The American Society for Microbiology (ASM) and Molecular and Cellular Biology (MCB) are issuing this Expression of Concern to alert readers to doubts about the integrity of the data in papers coauthored by Dr. Shigeaki Kato. In 2012, ASM was notified that the University of Tokyo Institute of Molecular and Cellular Biosciences is conducting an investigation of possible scientific misconduct by Dr. Kato. Among the publications under investigation for possible data manipulation are these five MCB articles. The Expression of Concern is only for papers coauthored by Dr. Kato that have been published since 2007, in accordance with the DHHS/ORI six-year limitation on research misconduct (http://ori.dhhs.gov/sites/default/files/42_cfr_parts_50_and_93_2005.pdf). MCB has contacted the Research Promotion Department of the University of Tokyo and asked to be informed of the findings of their ongoing investigation. Once ASM has been notified of the outcome of the investigation by the University of Tokyo, MCB will take appropriate action regarding these publications.

An airway-selective DNase hypersensitive site (DHS) at -35kb 5' to the cystic fibrosis transmembrane conductance regulator (CFTR) gene is evident in many lung cell lines and primary human tracheal epithelial cells but is absent from intestinal epithelia. DHS-35kb contains an element with enhancer activity in 16HBE14o- airway epithelial cells and is enriched for mono-methylated H3K4 histones (H3K4me1). We now define a 350 bp region within DHS-35kb, which has full enhancer activity and binds interferon regulatory factor-1 (IRF1) and nuclear factor Y (NF-Y) in vitro and in vivo. SiRNA-mediated depletion of IRF1 or overexpression of IRF2, an antagonist of IRF1, reduces CFTR expression in 16HBE14o- cells. NF-Y is critical for maintenance of H3K4me1 enrichment at DHS-35kb since depletion of NF-YA, a subunit of NF-Y, reduces H3K4me1 enrichment at this site. Moreover, depletion of SETD7, an H3K4 monomethyl transferase, reduces both H3K4me1 and NF-Y occupancy suggesting a requirement of H3K4me1 for NF-Y binding. NFY depletion also represses Sin3A and reduces its occupancy across the CFTR locus, which is accompanied by an increase in p300 enrichment at multiple sites. Our results reveal that the DHS-35kb airway selective enhancer element plays a pivotal role in regulation of CFTR expression by two independent regulatory mechanisms.

Androgens play a major role in the regulation of normal ovarian function, however, they are also involved in the development of ovarian pathologies. These contrasting effects may involve a differential response of granulosa cells to the androgens, testosterone (T) and dihydrotestosterone (DHT). To determine the molecular pathways that mediate the distinct effects of T and DHT, we studied the expression of liver receptor homolog-1 (LRH-1), a gene differentially regulated by these steroids. We found that although both T and DHT stimulate androgen receptor (AR) binding to the LRH-1 promoter, DHT prevents T mediated stimulation of LRH-1 expression. T stimulated the expression of aryl hydrocarbon receptor (AHR) and its interaction with the AR. T also promoted the recruitment of the AR/AHR complex to the LRH-1 promoter. These effects were not mimicked by DHT. We also observed that the activation of extracellular regulated kinases by T is required for AR and AHR interaction. In summary, T, but not DHT, stimulates AHR expression and the interaction between AHR and AR leading to the stimulation of LRH-1 expression. These findings could explain the distinct response of granulosa cells to T and DHT and provide a molecular mechanism by which DHT negatively affects ovarian function.

In contrast to prokaryotes, the precise mechanism of incorporation of ribosomal proteins into ribosomes in eukaryotes is not well understood. For the majority of eukaryotic ribosomal proteins, residues critical for rRNA binding, a key step in the hierarchical assembly of ribosomes, have not been well defined. In this study, we used the mammalian ribosomal protein L13a as a model to investigate the mechanism(s) underlying eukaryotic ribosomal protein incorporation into ribosomes. This work identified the Arginine residue at position 68 of L13a as essential for L13a binding to rRNA and incorporation into ribosomes. We also demonstrated that incorporation of L13a takes place during maturation of the 90S pre-ribosome in the nucleolus, but that translocation of L13a into the nucleolus is not sufficient for its incorporation into ribosomes. Incorporation of L13a into the 90S pre-ribosome was required for rRNA methylation within the 90S complex. However, mutations abolishing ribosomal incorporation of L13a did not affect its ability to be phosphorylated or its extra-ribosomal function in GAIT element-mediated translational silencing. These results provide new insights into the mechanism of ribosomal incorporation of L13a and will be useful in guiding future studies aimed at fully deciphering mammalian ribosome biogenesis.

The involvement of nuclear factor-kappaB (NF-κB) in several processes in the postnatal and adult brain, ranging from neuronal survival to synaptogenesis and plasticity, has been documented. In contrast, little is known about the functions of NF-κB during embryonic brain development. It is shown here that NF-κB is selectively activated in neocortical neural progenitor cells in the developing mouse telencephalon. Blockade of NF-κB activity leads to premature cortical neuronal differentiation and depletion of the progenitor cell pool. Conversely, NF-κB activation causes decreased cortical neurogenesis and expansion of the progenitor cell compartment. This effect is antagonized by the pro-neuronal transcription factor Hes6, which physically and functionally interacts with RelA-containing NF-κB complexes in cortical progenitor cells. In turn, NF-κB exerts an inhibitory effect on the ability of Hes6 to promote cortical neuronal differentiation. These results reveal previously uncharacterized functions, and modes of regulation, for NF-κB and Hes6 during cortical neurogenesis.

The sphingolipid backbone ceramide (Cer) is a major component of lipid lamellae in the stratum corneum of epidermis and has a pivotal role in epidermal barrier formation. Unlike Cers in other tissues, Cers in epidermis contain extremely long fatty acids (FAs). Decreases in epidermal Cer levels, as well as changes in their FA chain-lengths, cause several cutaneous disorders. However, the molecular mechanisms that produce such extremely long Cers and determine their chain-lengths are poorly understood. We generated mice deficient in the Elovl1 gene, encoding the FA elongase responsible for producing C20-28 FAs. The Elovl1 knockout mice died shortly after birth due to epidermal barrier defects. The lipid lamellae in the stratum corneum were largely diminished in these mice. In the epidermis of the Elovl1-null mice, the levels of Cers with ≥C26 FAs were decreased, while those of Cers with ≤C24 FAs were increased. In contrast, the levels of C24 sphingomyelin were reduced, accompanied by an increase in C20 sphingomyelin levels. Two ceramide synthases, CerS2 and CerS3, expressed in an epidermal layer-specific manner, regulate Elovl1 to produce acyl-CoAs with different chain-lengths. Elovl1 is a key determinant of epidermal Cer chain-length and essential for permeability barrier formation.

Net1 is a RhoA GEF that is overexpressed in a subset of human cancers and contributes to cancer cell motility and invasion in vitro. However, the molecular mechanism accounting for its role in cell motility and invasion has not been described. In the present work we show that expression of both Net1 isoforms in breast cancer cells is required for efficient cell motility. Although loss of Net1 isoform expression only partially blocks RhoA activation, it inhibits LPA-stimulated migration as efficiently as knockdown of RhoA itself. However, we demonstrate that the Net1A isoform predominantly controls myosin light chain phosphorylation and is required for trailing edge retraction during migration. Net1A interacts with FAK, localizes to focal adhesions, and is necessary for FAK activation and focal adhesion maturation during cell spreading. Net1A expression is also required for efficient invasion through a Matrigel matrix. Analysis of invading cells demonstrates that Net1A is required for amoeboid-type invasion and loss of Net1A expression causes cells to shift to a mesenchymal phenotype characterized by high β1-integrin activity and MT1-MMP expression. These results demonstrate a previously unrecognized role for the Net1A isoform in controlling FAK activation during planar cell movement and amoeboid motility during ECM invasion.

IL-1β is a potent proinflammatory and immune-regulatory cytokine playing an important role in the progression of rheumatoid arthritis (RA). However, the signaling network of IL-1β in synoviocytes from RA is still poorly understood. Here, we show for the first time that phospholipase D1 (PLD1) but not PLD2, is selectively upregulated in the IL-1β-stimulated synoviocytes as well as synovium from RA patients. IL-1β enhanced the binding of NFκB and ATF-2 to PLD1 promoter, thereby enhancing PLD1 expression. PLD1 inhibition abolished IL-1β-induced expression of proinflammatory mediators and angiogenic factors by suppressing the binding of NFκB or HIF-1α to the promoter of its target genes as well as IL-1β-induced proliferation or migration. However, suppression of PLD1 activity promoted cell cycle arrest via transactivation of FoxO3a. Furthermore, PLD1 inhibitor significantly suppressed joint inflammation and destruction in IL-1 receptor antagonist deficient mice (IL-1Ra-/-), a model of spontaneous arthritis. Taken together, these results suggest that the abnormal upregulation of PLD1 may contribute to the pathogenesis of IL-1β-induced chronic arthritis, and a selective PLD1 inhibitor might provide a potential therapeutic molecule for the treatment of chronic inflammatory autoimmune disorders.