 Hello, I am Maria Chiaroscaini and I am a researcher of the Immunology and Molecular Oncology Unit at the Veneto Institute of Oncology in Padua, in Italy. Our melanoma group is part of a much larger multidisciplinary regional melanoma network, a result of a joint effort between the Veneto Institute of Oncology and the University of Padua. And it is also part of the Italian Melanoma Intergroup, a no-profit organization from Epidemiology Prevention Diagnosis and Treatment of Malignal Melanoma. I'd like to introduce you today to our article recently published in Human Mutation entitled CDK N2A Unclassified Variants in Familial Malignal Melanoma, combining functional and computational approaches for their assessment. This work was made possible through a strict collaboration between our unit and the Department of Biomedical Sciences of the University of Padua. CDK N2A is the most common highly penetrant susceptibility gene identified today in Familial Malignal Melanoma. It codes for P16 and Ongosupressor involved in sacroagulation. A growing number of unclassified variants of this protein are being identified. Moreover, germline P16 mutations are found in about 40% of melanoma-prone families. Anyway, unless the pathogenic role of this variant can be demonstrated, they cannot be used for genetic counseling of at-risk individuals in melanoma-prone families. In our work, we try to assess the role of P16 unclassified variants in melanoma-on-supressor and propose new tools for P16 variants classification. We set up a protocol combining computational and experimental tools in an integrated approach. We analyzed both the effects on the P16 ability of controlling cell cycle transition and the effects on protein structure level with particular emphasis on protein stability and ability to bind its targets. Its peculiar structure, composed of four anchoring domains, has been deeply investigated. In doing this, we selected to putogenize to all the possible amino acid variants two highly-conserved P16 glycine positions. Along with these variants, also a few variants from melanoma patients were analyzed. The behavior of different amino acids at the same protein position has been investigated to shed some light on different aspects as relevance and permissiveness for amino acid replacement of different P16 sites and importance of the specific type of substitution. The relationship between a specific amino acid change and the chemical nature of the involved interaction was also investigated. For the in-vit approach, P16 variants obtained by cell-specific mutagenesis were expressed in a P16 cell line and tested for the ability to block cell proliferation. In parallel, these variants underwent in silico-prediction analysis and molecular dynamics simulations, as you can see now in this video, representing a 10 nanoseconds dynamics of P16 bound to its target CDH6. We predicted whether the structural effects cause unfolding or local instability, which might interfere with protein-protein interactions. Merging of the in-vitoid in silico data should solve the variants' classification problem by providing tools for the interpretation of the structural and functional impact of P16 variants, while also providing an explanation of why a specific variant displays a specific behavior or why an apparently innocent institution could cause a mutation-like phenotype. In conclusion, this approach could represent a pilot study for the definition of a protocol applicable to other unclassified variants involved in other diseases as well. I would like to thank all the people involved in this work and I hope I've inspired you in reading the full article published in Human Mutation Journal. And if you have any questions or comments, please don't hesitate to contact me at the email address shown below. Thank you for your attention.