MY01, a class XIV myosin, affects developmentally‐regulated elimination of the macronucleus during conjugation of Tetrahymena thermophila

Abstract

Background information. Nuclear dimorphism is characteristic of ciliated protozoa. A transcriptionally-active macronucleus co-exists with a transcriptionally-silent micronucleus, which is activated only at conjugation. During conjugation, each conjugant develops two new genetically matched macronuclei and micronuclei, and the pre-existing macronucleus is eliminated. Elimination of the pre-existing macronucleus during conjugation is an apoptotic-like process. The macronucleus becomes highly condensed, DNA laddering occurs, caspase activity increases, acidic enzymes accumulate within the nucleoplasm, and the nucleus shrinks in size. The current study focused on the involvement of actin and myosin in nuclear events of conjugation. A myosin knockout strain was mated with wild-type, and the nuclear events were monitored with confocal microscopy.Results. Early nuclear events, including development of new macronuclei and micronuclei, appeared qualitatively normal in knockout conjugants. Completion of nuclear condensation and acidification in the pre-existing macronucleus was blocked in 44% of knockout conjugants. Knockout conjugants that failed to fully achieve nuclear condensation and acidification did not eliminate the pre-existing macronucleus. In control experiments, blockage of chromatin condensation, nuclear acidification, and macronuclear elimination was never observed in wild-type conjugants.Conclusions. Perturbation of either DNA fragmentation, chromatin condensation or nuclear acidification can lead to blockage of apoptotic-like elimination of the macronucleus in MYO1-knockout conjugants. Consistent with the known motor function of myosins and the involvement of Myo1 in vesicle trafficking in Tetrahymena, we argue that Myo1 could specifically affect condensation of chromatin and acidification of the nucleus through direct interaction with chromatin and through Myo1-dependent vesicle trafficking to the nucleus.

Identification, modeling, and characterization studies ofTetrahymena thermophilamyosinFERMdomains suggests a conserved core fold but functional differences

“…We propose that T. thermophila myosin FERM possess the ability to bind various phosphoinositides (PIs) and bind with higher affinity to monophosphorylated PIs, suggesting divergent functionality from canonical FERM domains. These results are in line with experimental studies which also suggest divergent functional traits for T. thermophila myosins [Garcés et al, ; Gotesman et al, ]. The results presented here lay a foundation for further experimental investigation specifically of T. thermophila myosin FERM domains and also provide insight into the functional role of the FERM domain in general.…”

A FERM domain in a class XIV myosin interacts with actin and tubulin and localizes to the cytoskeleton, phagosomes, and nucleus in Tetrahymena thermophila

“…Amitosis of the macronucleus is frequently aberrant in the knockout due to failure of nuclear elongation, which results in unequal segregation of macronuclear DNA. In the knockout, developmentally regulated destruction of the macronucleus fails to occur during conjugation [Garcés et al, 2009]. How Myo1 recognizes its localization targets is unknown.…”

Unique sequences and predicted functions of myosins in Tetrahymena thermophila

“…The cellular function of Myo1 has only been analyzed by one research group: that is, Gavin and colleagues at Brooklyn College of the City, University of New York. MYO1 knockout cells fail to form and transport phagosomes, to elongate the macronucleus and segregate macronuclear DNA during cell division, and to condense chromatin and acidify the old macronucleus during the destruction of the old macronucleus during mating (Williams et al, 2000;Hosein et al, 2005;Hosein and Gavin, 2007;Garcés et al, 2009). A myosin that forms a dimer and contains a long fibrous tail was purified from T. pyriformis (Kanzawa et al, 1996).…”