Doneddu, Valentina (2014) Med1 /MBD4 DNA repair enzyme and SUMOylation. Doctoral Thesis.
Different and recent studies have shown that the post-translational modification, called SUMOylation, plays a central role about the function, stability, localization and activity of the proteins. The SUMOylation regulates different cellular processes, including transcription, replication,chromosome segregation and DNA repair. The SUMO proteins share the structural conformation with the Ubiquitin family proteins. The SUMOylation cycle starts when the activating enzyme E1 recognizes the mature isoform of SUMO and forms a complex with it. After that, the conjugating enzyme E2 drives the SUMO to the protein target. Sometimes this final step, requires the presence of ligases E3. Now are known 4 isoforms of SUMO :SUMO1-4. SUMO2 and SUMO3 have 97% identical aminoacid sequence and for this reason are commonly referred as SUMO2/3. SUMO1 shares just 50% aminoacid sequence with SUMO2/3. The most characterized isoforms are SUMO1 and SUMO2/3.SUMO4 is 87% similar to SUMO2 but is unclear how is processed and conjugated. SUMO4 is just present in the liver and in the brain. By contrast, SUMO1 and SUMO2/3 are ubiquitously expressed. Human thymidine DNA glycosylase, hTDG, is a monofunctional DNA glycosylase that excises thymine and uracil from G-T and G-U mismatched oligonucleotide. This kind of DNA repair system is known as BER: base excision repair. Other enzymes that work in the BER system are MED 1/MBD4. In the 2002, Primo Schar shows that hTDG , when sumoylated, is facilitated in its enzymatic turnover. After that, the SUMOylation involved in the DNA repair system has become object of speculation, considering also the strong link between DNA repair and cancer. SUMOylation is a post-translational modification characterized by covalent and reversible binding of small ubiquitin-like modifier (SUMO) to a target protein . The four isoforms of SUMO, termed SUMO-1, -2, -3, are ubiquitously expressed, unlike SUMO -4 which is tissue specific. In general, SUMO, can alter the stability, localization and activity of protein targets. SUMO is also involved in cellular processes such as mitosis, cell development and differentiation, senescence and apoptosis. SUMOylation occurs also in proteins involved in the mechanisms of DNA repair (Smet-Nocca et al 2011). In order to investigate the possibility that MED1, the base excision repair (BER) enzyme (Bellacosa et al.1999), is SUMOylated, we have performed Co-Immunoprecipitation and Western blot analyses . We focused our attention on the presence of consensus sites of SUMOylation and we have also studied MED1 in response to DNA damage reagents, when MED1 is mutated in these sites.
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