Discovery of a Family of Genomic Sequences Which Interact Specifically with the c-MYC Promoter to Regulate c-MYC Expression

Abstract

G-quadruplex forming sequences are particularly enriched in the promoter regions of eukaryotic genes, especially of oncogenes. One of the most well studied G-quadruplex forming sequences is located in the nuclease hypersensitive element (NHE) III1 of the c-MYC promoter region. The oncoprotein c-MYC regulates a large array of genes which play important roles in growth regulation and metabolism. It is dysregulated in >70% of human cancers. The silencer NHEIII1 located upstream of the P1 promoter regulates up-to 80% of c-MYC transcription and includes a G-quadruplex structure (Pu27) that is required for promoter inhibition. We have identified, for the first time, a family of seventeen G-quadruplex-forming motifs with >90% identity with Pu27, located on different chromosomes throughout the human genome, some found near or within genes involved in stem cell maintenance or neural cell development. Notably, all members of the Pu27 family interact specifically with NHEIII1 sequence, in vitro. Crosslinking studies demonstrate that Pu27 oligonucleotide binds specifically to the C-rich strand of the NHEIII1 resulting in the G-quadruplex structure stabilization. Pu27 homologous sequences (Pu27-HS) significantly inhibit leukemic cell lines proliferation in culture. Exposure of U937 cells to the Pu27-HS induces cell growth inhibition associated with cell cycle arrest that is most likely due to downregulation of c-MYC expression at the RNA and/or protein levels. Expression of SOX2, another gene containing a Pu27-HS, was affected by Pu27-HS treatment as well. Our data suggest that the oligonucleotides encoding the Pu27 family target complementary DNA sequences in the genome, including those of the c-MYC and SOX2 promoters. This effect is most likely cell type and cell growth condition dependent. The presence of genomic G-quadruplex-forming sequences homologous to Pu27 of c-MYC silencer and the fact that they interact specifically with the parent sequence suggest a common regulatory mechanism for genes whose promoters contain these sequences.

DNA Conformation Regulates Gene Expression: The MYC Promoter and Beyond

“…Thousands of sequences with predicted capacity to form G‐quadruplexes have been identified throughout the genome. Using oligonucleotide probes, sequences similar to the MYC G‐quadruplex were observed at multiple promoters, indicating broader regulatory roles in transcription . Moreover, ChIP‐seq analysis using an antibody that recognizes G‐quadruplex structure revealed enrichment of peaks at nucleosome‐depleted regions .…”

“…G‐quadruplex structures can also influence promoter‐proximal Pol II pausing. For example, stabilization of the G‐quadruplex, by treatment of lymphoma cell lines with oligonucleotides targeting the C‐rich strand, results in reduced proliferation and decreased MYC expression . The i‐motif structure was originally characterized in vitro, due to the highly acidic conditions required for formation .…”

“…For example, stabilization of the G-quadruplex, by treatment of lymphoma cell lines with oligonucleotides targeting the C-rich strand, results in reduced proliferation and decreased MYC expression. [85] The imotif structure was originally characterized in vitro, due to the highly acidic conditions required for formation. [80] However, in the presence of negative supercoiling, stabilization of the i-motif is possible under physiological pH, further highlighting the importance of torsional stress to DNA conformation.…”

“…Using oligonucleotide probes, sequences similar to the MYC Gquadruplex were observed at multiple promoters, indicating broader regulatory roles in transcription. [85] Moreover, ChIP-seq analysis using an antibody that recognizes G-quadruplex structure revealed enrichment of peaks at nucleosome-depleted regions. [6] Interestingly, the sequences with predicted capacity to form G-quadruplexes are particularly enriched at promoters of oncogenes, while tumor suppressor promoters are underrepresented, indicating that gene function is linked with local genomic structure.…”

The Frequency of Double Expresser in Selected Cases of High Grade Diffuse Large B-Cell Lymphomas

“…After gene rearrangements for immunoglobulins and expression of specific surface markers, the mature naïve B-cells leave the bone marrow and settles into the lymphoid tissue (Young et al, 1999). c-Myc is known to be a "master regulator" of the genes that are involved in metabolism and cell cycle growth initiating and maintaining molecular mechanisms (Rezzoug et al, 2016). It is a transcription factor located on chromosome 8q24 that is crucial for early B-cell development in the bone marrow by activating a transcription factor required for the maintenance of B-cells identity (Nguyen et al, 2017).…”