Letter to the Editor

Trans-silencing by P Elements Inserted in Subtelomeric Heterochromatin Involves the Drosophila Polycomb Group Gene, Enhancer of Zeste

Corresponding author: Donald C. Rio, 401 Barker Hall, University of California, Berkeley, CA 94720-3204., don_rio{at}uclink4.berkely.edu (E-mail)

REGARDING the article that Siobhan Roche and I published in the August 1998 issue of GENETICS (vol. 149, pp. 1839–1855): In the course of trying to extend the observations in the article, we have found that we cannot reproduce the results concerning the role of the polycomb group gene, Enhancer of zeste [E(z)], in the trans-silencing of P-element transgenes by P elements in subtelomeric heterochromatin. [We have not retested the effect of the E(z) alleles on repression of gonadal dysgenic (GD) sterility.] We have spent the past 6 months doing controls, repeating crosses and checking, remaking, or getting new copies of stocks used in these studies. Outlined below, in detail, is a summary of our findings since the article was published.

In our original article, seven different alleles of E(z) were tested for their effects on trans-silencing of P-element transgenes. We observed a loss of trans-silencing in the presence of four different E(z) alleles.

1. The original observation that E(z) is required for the trans-silencing of P-element transgenes was made with the E(z)⁶¹ allele. For this experiment, the E(z)⁶¹ allele (which is marked with the recessive ebony mutation) was crossed into an Lk-P(1A);+/TM3 background and black-bodied progeny were selected. The presence of the Lk-P(1A) P elements was confirmed by DNA blot hybridization (Southern blotting). The E(z)⁶¹ allele no longer affects the trans-silencing of an hsp83-IVS3-LacZ reporter transgene. The assumption now is that there was originally a modifier mutation present in the stock that contributed to the loss of trans-silencing and that has since (over the past 2 years) been lost from the stock.

2. The second set of data in which we observed a loss of trans-silencing involved three additional E(z) alleles [E(z)²⁸, E(z)³², and E(z)⁶⁰]. For these tests, the E(z) alleles were crossed into an Lk-P(1A);CxD/TM3 background. The Lk-P(1A);CxD/TM3 strain was generated by crossing the CxD mutation in to the Lk-P(1A);+/TM3 background and was shown to repress P-element transposition as effectively as the Lk-P(1A);+/TM3 strain. We first used the Lk-P(1A);CxD/TM3 strain to test the effect of the polycomb allele, Pc¹⁶, on trans-silencing and did not observe a derepression of the reporter transgene. However, the three additional E(z) alleles that affected trans-silencing were also tested in the Lk-P(1A);CxD/TM3 background. We have tried unsuccessfully for the last year to repeat these results and made the unfortunate discovery that over time, the Lk-P(1A) P elements became unstable in the Lk-P(1A);CxD/TM3 strain and its E(z)^- derivatives. This effect seems to be peculiar to this stock, since normally the Lk-P(1A) P elements are extremely stable. While we had tested for the presence of the Lk-P(1A) P elements in the Lk-P(1A); E(z)^-/TM3 strains by Southern blotting at the time the genetic experiments were performed, subsequent PCR analysis of single flies from samples frozen at that time (8/96) indicated that approximately half of the individuals in the population had lost the Lk-P(1A) P elements. This P-element instability explains the apparent loss of trans-silencing that was observed in 1995–96 and reported in the article.

3. Finally, the reported observation that P-element enhancer traps at 1A or 100F also exhibit trans-silencing is correct, but the observation that the E(z)³² allele reduces this effect cannot be repeated. We know that our inability to repeat these results cannot be due to instability of the P-element enhancer traps but may also be a result of background modifiers or changes in the strength of the trans-silencing, which we had observed before.

This article has been cited by other articles:

				M. J. Simmons, D.-F. Ryzek, C. Lamour, J. W. Goodman, N. E. Kummer, and P. J. Merriman Cytotype Regulation by Telomeric P Elements in Drosophila melanogaster: Evidence for Involvement of an RNA Interference Gene Genetics, August 1, 2007; 176(4): 1945 - 1955. [Abstract] [Full Text] [PDF]

				K. J. Haley, J. R. Stuart, J. D. Raymond, J. B. Niemi, and M. J. Simmons Impairment of Cytotype Regulation of P-Element Activity in Drosophila melanogaster by Mutations in the Su(var)205 Gene Genetics, October 1, 2005; 171(2): 583 - 595. [Abstract] [Full Text] [PDF]