The mdml mutation causes temperature-sensitive growth and defective transfer of nuclei and mitochondria into developing buds of yeast cells at the nonpermissive temperature. The MDM1 gene was cloned by complementation, and its sequence revealed an open reading frame encoding a potential protein product of 51.5 kD. This protein displays amino acid sequence similarities to hamster vimentin and mouse epidermal keratin. Gene disruption demonstrated that MDM1 is essential for mitotic growth. Antibodies against the MDM1 protein recognized a 51-kD polypeptide that was localized by indirect immunofluorescence to a novel pattern of spots and punctate arrays distributed throughout the yeast cell cytoplasm. These structures disappeared after shifting mdm1 mutant cells to the nonpermissive temperature, although the cellular level of MDM1 protein was unchanged. Affinity-purified antibodies against MDM1 also specifically recognized intermediate filaments by indirect immunofluorescence of animal cells. These results suggest that novel cytoplasmic structures containing the MDM1 protein mediate organelle inheritance in yeast.