Aberrant expression of sialic acid containing glycolipids, or gangliosides and their corresponding synthases have been associated with a variety of cancers. GM2-synthase,the enzyme responsible for the formation of a pro-tumorigenic ganglioside, GM2 is well reported to be overexpressed across various cancer tissues and cell lines. This over-expression of GM2-synthase has been linked with increased migration, invasion and epithelial to mesenchymal transition (EMT) as well as induction of a local and systemic host immune suppression in cancer. Despite only a handful of studies demonstrating an epigenetic mechanism underlying the transcriptional regulation of GM2-synthase gene, the detailed mechanism still remains unclear. Here we identified the total proteome associated with the GM2-synthase promoter through a modified CRISPR-dCas9 based proteome profiling approach by categorizing all the identified proteins leading to a detailed elucidation of the molecular drivers behind GM2-synthase transcription. While the previous study identified an acetylation-dependent de-repression of the transcription factor SP1 causing GM2-synthase activation, the underlying molecular mechanism driving its activation wasn’t clear. This study demonstrated that the histone acetyltransferase (HAT), p300 acts a pivotal factor which on one hand cause acetylation-mediated degradation of SP1, and on the other hand modulates SMAD2/4 to have a direct positive impact on GM2 synthase gene transcription. We identified p300 to have an activator role in GM2 synthase gene transcription through knock out, knock down and over-expression experiments. Furthermore, SP1 degradation, SMAD activation and their DNA binding patterns shows the important role of p300 on SP1 and SMAD complexes. Altogether we have identified SMAD2/4 as an activator complex, p300 as a positive regulator and uncovered a critical p300-SMAD-SP1 regulatory axis in GM2--synthase transcriptional regulation.