SWIM: A computational tool to unveiling crucial nodes in complex biological networks

SWItchMiner (SWIM) is a wizard-like software implementation of a procedure, previously described, able to extract information contained in complex networks. Specifically, SWIM allows unearthing the existence of a new class of hubs, called "fight-club hubs", characterized by a marked negative correlation with their first nearest neighbors. Among them, a special subset of genes, called "switch genes", appears to be characterized by an unusual pattern of intra- and inter-module connections that confers them a crucial topological role, interestingly mirrored by the evidence of their clinic-biological relevance. Here, we applied SWIM to a large panel of cancer datasets from The Cancer Genome Atlas, in order to highlight switch genes that could be critically associated with the drastic changes in the physiological state of cells or tissues induced by the cancer development. We discovered that switch genes are found in all cancers we studied and they encompass protein coding genes and non-coding RNAs, recovering many known key cancer players but also many new potential biomarkers not yet characterized in cancer context. Furthermore, SWIM is amenable to detect switch genes in different organisms and cell conditions, with the potential to uncover important players in biologically relevant scenarios, including but not limited to human cancer

Shmt2: a stat3 signaling new player in prostate cancer energy metabolism

Prostate cancer (PCa) is a multifactorial disease characterized by the aberrant activity of different regulatory pathways. STAT3 protein mediates some of these pathways and its activation is implicated in the modulation of several metabolic enzymes. A bioinformatic analysis indicated a STAT3 binding site in the upstream region of SHMT2 gene. We demonstrated that in LNCaP, PCa cells' SHMT2 expression is upregulated by the JAK2/STAT3 canonical pathway upon IL-6 stimulation. Activation of SHTM2 leads to a decrease in serine levels, pushing PKM2 towards the nuclear compartment where it can activate STAT3 in a non-canonical fashion that in turn promotes a transient shift toward anaerobic metabolism. These results were also confirmed on FFPE prostate tissue sections at different Gleason scores. STAT3/SHMT2/PKM2 loop in LNCaP cells can modulate a metabolic shift in response to inflammation at early stages of cancer progression, whereas a non-canonical STAT3 activation involving the STAT3/HIF-1α/PKM2 loop is responsible for the maintenance of Warburg effect distinctive of more aggressive PCa cells. Chronic inflammation might thus prime the transition of PCa cells towards more advanced stages, and SHMT2 could represent a missing factor to further understand the molecular mechanisms responsible for the transition of prostate cancer towards a more aggressive phenotyp

The laminA/NF-Y protein complex reveals an unknown transcriptional mechanism on cell proliferation

Lamin A is a component of the nuclear matrix that also controls proliferation by largely unknown mechanisms. NF-Y is a ubiquitous protein involved in cell proliferation composed of three subunits (-YA -YB -YC) all required for the DNA binding and transactivation activity. To get clues on new NF-Y partner(s) we performed a mass spectrometry screening of proteins that co-precipitate with the regulatory subunit of the complex, NF-YA. By this screening we identified lamin A as a novel putative NF-Y interactor. Co-immunoprecipitation experiments and confocal analysis confirmed the interaction between the two endogenous proteins. Interestingly, this association occurs on euchromatin regions, too. ChIP experiments demonstrate lamin A enrichment in several promoter regions of cell cycle related genes in a NF-Y dependent manner. Gain and loss of function experiments reveal that lamin A counteracts NF-Y transcriptional activity. Taking advantage of a recently generated transgenic reporter mouse, called MITO-Luc, in which an NF-Y–dependent promoter controls luciferase expression, we demonstrate that lamin A counteracts NF-Y transcriptional activity not only in culture cells but also in living animals. Altogether, our data demonstrate the occurrence of lamin A/NF-Y interaction and suggest a possible role of this protein complex in regulation of NF-Y function in cell proliferatio

NF-Y Dependent Epigenetic Modifications Discriminate between Proliferating and Postmitotic Tissue

The regulation of gene transcription requires posttranslational modifications of histones that, in concert with chromatin remodeling factors, shape the structure of chromatin. It is currently under intense investigation how this structure is modulated, in particular in the context of proliferation and differentiation. Compelling evidence suggests that the transcription factor NF-Y acts as a master regulator of cell cycle progression, activating the transcription of many cell cycle regulatory genes. However, the underlying molecular mechanisms are not yet completely understood. Here we show that NF-Y exerts its effect on transcription through the modulation of the histone “code”. NF-Y colocalizes with nascent RNA, while RNA polymerase II is I phosphorylated on serine 2 of the YSPTSPS repeats within its carboxyterminal domain and histones are carrying modifications that represent activation signals of gene expression (H3K9ac and PAN-H4ac). Comparing postmitotic muscle tissue from normal mice and proliferating muscles from mdx mice, we demonstrate by chromatin immunoprecipitation (ChIP) that NF-Y DNA binding activity correlates with the accumulation of acetylated histones H3 and H4 on promoters of key cell cycle regulatory genes, and with their active transcription. Accordingly, p300 is recruited onto the chromatin of NF-Y target genes in a NF-Y-dependent manner, as demonstrated by Re-ChIP. Conversely, the loss of NF-Y binding correlates with a decrease of acetylated histones, the recruitment of HDAC1, and a repressed heterochromatic state with enrichment of histones carrying modifications known to mediate silencing of gene expression (H3K9me3, H3K27me2 and H4K20me3). As a consequence, NF-Y target genes are downregulated in this context. In conclusion, our data indicate a role of NF-Y in modulating the structure and transcriptional competence of chromatin in vivo and support a model in which NF-Y-dependent histone “code” changes contribute to the proper discrimination between proliferating and postmitotic cells in vivo and in vitro

Requirement for Down-Regulation of the CCAAT-binding Activity of the NF-Y Transcription Factor during Skeletal Muscle Differentiation

NF-Y is composed of three subunits, NF-YA, NF-YB, and NF-YC, all required for DNA binding. All subunits are expressed in proliferating skeletal muscle cells, whereas NF-YA alone is undetectable in terminally differentiated cells in vitro. By immunohistochemistry, we show that the NF-YA protein is not expressed in the nuclei of skeletal and cardiac muscle cells in vivo. By chromatin immunoprecipitation experiments, we demonstrate herein that NF-Y does not bind to the CCAAT boxes of target promoters in differentiated muscle cells. Consistent with this, the activity of these promoters is down-regulated in differentiated muscle cells. Finally, forced expression of the NF-YA protein in cells committed to differentiate leads to an impairment in the down-regulation of cyclin A, cyclin B1, and cdk1 expression and is accompanied by a delay in myogenin expression. Thus, our results indicate that the suppression of NF-Y function is of crucial importance for the inhibition of several cell cycle genes and the induction of the early muscle-specific program in postmitotic muscle cells