Two different promoters were used in luciferase assays to test this hypothesis: the BAX promoter that regulates the expression of Bax and the MDM2 promoter that regulates the expression of the Mdm2 protein, a potent inhibitor of p53 Albaspidin-AA transcriptional activation. Ankrd2 enhanced the p53-mediated activation of both the BAX and MDM2 promoters, while ZASP6 acted as a negative regulator of p53 transcriptional activation both in the presence and absence of Ankrd2. Further experimental evidence is needed in order to elucidate the possible influence of ZASP and Ankrd2 on p53 function. The question is when and why ZASP acts to a enuate p53 transactivation and how Ankrd2 is involved. Since. ZASP can interact with p53 preventing it to activate p53 responsive genes, ZASP can also be involved in mechanoptosis, similarly to what is proposed for telethonin. Mechanoptosis is defined as a type of apoptosis induced via mechanical stress. The increase in sarcomere activity is translated into pro-survival signals by sequestering p53 in cytoplasm. Telethonin interferes with p53 in nucleus, but we can propose an additional mechanism of cytoplasmic sequestration of p53 when muscle cells are challenged and sarcomeroptosis is activated. Ankrd2 could also be sequestered thus preventing the activation of p53 in nucleus. The study of Lin and colleagues on PKA and ZASP directly connects Cypher/ZASP to the signal transduction pathways via participation in signaling that regulates the phosphorylation of some proteins, including the L-type Ca+ channel and Cypher/ZASP itself. In cardiomyocytes they show that not only does Cypher/ZASP interact with the regulatory subunit of PKA acting as physical anchor but that it was also phosphorylated by it at Ser265 and Ser296. Also the Cypher/ZASP PDZ domain enhanced PKA phosphorylation of LTCC protein by binding via its C-terminal PDZ binding motif. Another interesting fact was that they showed Cypher/ZASP binding to Coptisine-chloride calcineurin, a serine/threonine phosphatase of the LTCC. These and our findings connecting ZASP to the p53 pathway are in line with the concept that in cardiac Z-lines ZASP not only has a structural function but also a role in signaling. Other intriguing information in this regard is that ZASP can bind proteins of the FATZ/calsarcin/myozenin family that in turn are known to bind and regulate calcineurin. In conclusion, here we describe the expression pa ern and dynamic properties of ZASP isoforms and disease associated mutants of ZASP. Ankrd2 and p53 were identified as new binding partners of ZASP. Moreover, these three proteins are able to form a trimeric complex, potentially involved in signaling, regulation of gene expression and muscle differentiation. Endogenous polySUMOylation of p53 occurred but only in the triple complex in the presence of ZASP6. ZASP6 was shown to be a negative regulator of p53, at least for transactivation of the p53-responsive promoters MDM2 and BAX. On studying disease associated mutations in ZASP6 we found that two ZM-motif mutants, A165V and A171T, were not able to bind Ankrd2. This is the first indication of how mutant ZASP protein can differ from wild type and is particularly important since the A165V mutation is responsible for a well characterized autosomal dominant distal myopathy, zaspopathy.