A natural mild isolate of Zucchini yellow mosaic virus was found to contain a mutation in the helper component (HC-Pro) within a conserved motif, "CDNQLD", located 12 residues downstream from the "FRNK" motif involved in symptom severity. Introducing the mutation in an infectious cDNA clone of ZYMV resulted in an almost complete absence of symptoms, although viral accumulation was only partially reduced. The FRNK(X)(12)CDNQLD sequence might be part of a larger motif that is conserved in potyviruses and plays a role in symptomatology and/or silencing inhibition.