Analysis of mating type determination by transplantation of O macronuclear karyoplasm in Paramecium tetraurelia

Abstract

The odd (0) or even (E) mating type in Paramecium tetruurelia is determined during the first cell cycle after new macronuclear development. The present paper demonstrates that mating type E is irreversibly determined at the end of the first cell cycle. Direct evidence comes from transplanting 0 macronuclear karyoplasm containing 0-determining factor into E autogamous cells during a new postzygotic macronuclear development. Transplantation of 0 macronuclear karyoplasm into E autogamous cells at 7-8 hr after the origin of the macronucleus from a product of the synkaryon produces nearly 100% 0 mating type among the exautogamous cell lines but almost none 10-11 hr after the origin of the macronucleus (around the end of the first cell cycle). The macronuclear anlagen at the stage at which mating type E seems to be fixed contains about 20 times as much DNA as the vegetative G1 micronucleus. The 0-determining factor shifting E cells toward 0 mating type by transplanting 0 macronuclear karyoplasm is also produced by the newly developed macronucleus in an effective concentration at 10-11 hr after the sensitive period and produced at full levels by the third cell cycle. The level of 0 factor in the macronucleus then gradually declines with subsequent repeated rounds of DNA synthesis and is finally lost by the eighth cell cycle.

Microinjection of plasmid DNA encoding the A surface antigen of Paramecium tetraurelia restores the ability to regenerate a wild-type macronucleus.

“…The same pattern of nucleocytoplasmic determination in P. tetraurelia has also been described for mating-type determination of the B group of species in the P. aurelia complex by Sonneborn (15,18) and for discharge and nondischarge of trichocysts by Sonnebom and Schneller (19). Direct evidence for the production of an odd-determining mating-type nuclear factor by the young 0 macronucleus and for the existence of an even-determining cytoplasmic factor in cells during development of the new zygotic macronuclei has been obtained by microinjection experiments (8,11,12). It should be noted that the cytoplasmic factors specified by the macronucleus were almost exclusively found in cells during new macronuclear development in both mating-type determination and repair of the cytoplasmic defect in forming normal macronuclei in d48.…”

Epigenetic Mechanisms Affecting Macronuclear Development in Paramecium and Tetrahymena

“…Moreover he proved with appropriate crosses that all the O * individuals were mtD 51 / mtD 51 . His hypothesis then was that in the mtD 51 / mtD 32 genotype a new cytoplasmic state, which he called O * was sometimes produced and it allowed the 1:1 segregation to be seen. The O * cytoplasmic state was somewhat less stable than either O or E. In fact, crosses of O * ( mtD 51 / mtD 51 × E ( mtD 32 / mtD 32 ) gave close to 100% segregants in the F‐2 by autogamy.…”

“…How does one explain these results? Brygoo suggested that 32 differed from 51 in a single gene at the mtD locus, the two stocks being mtD 32 / mtD 32 and mtD 51 / mtD 51 . He thought that in a cross of the two stocks, a true genie segregation occurred in the F‐2, but that it was mostly hidden by the cytoplasmic state of the cells.…”

“…His hypothesis then was that in the mtD 51 / mtD 32 genotype a new cytoplasmic state, which he called O * was sometimes produced and it allowed the 1:1 segregation to be seen. The O * cytoplasmic state was somewhat less stable than either O or E. In fact, crosses of O * ( mtD 51 / mtD 51 × E ( mtD 32 / mtD 32 ) gave close to 100% segregants in the F‐2 by autogamy. An appropriate series of crosses revealed that the mtD locus was different (Brygoo 1977) from the mtA °, mtB °, and mtC ° loci identified by Byrne.…”

Mating-Type Inheritance

“…Direct evidence of cytoplasmic E-determining factors has been obtained by transplanting E cytoplasm into 0 cells just before development of the new macronucleus, which induced transformation of o to E in the progeny clones. Such O-determining "karyoplasmic" factors may be produced or activated at conjugation and then decrease in activity in subsequent fissions after the development of a new macronucleus (Koizumi and Kobayashi 1984). However, when the 0 macronucleus was transplanted into the cytoplasm of E cells, E ex autogamous clones were transformed to O.…”