OF DROSOPHILA MELANOGASTER by gyvwpsjkko

VIEWS: 26 PAGES: 11

									                  POLYTENIZATION OF THE RIBOSOMAL GENES
                      ON THE X AND Y CHROMOSOMES
                                   OF
                       DROSOPHILA MELANOGASTER

                                       SHARYN A. ENDOW
        Department of Microbiology and Immunology, Duke University Medical Center,
                               Durham, North Carolina 27720
                                  Manuscript received July 23, 1981
                               Revised copy accepted December 22, 1981

                                             ABSTRACT

              It has previously been shown (ENDOW      and GLOVER    1979), that polyteni-
          zation of the ribosomal genes in D. melanogaster Ore-R X / Y cells and in
          hybrid X / X cells (ENDOW    1980) involves replication of genes predominantly
          from one of the cell’s two nucleolus organizers. This analysis takes advantage
          of strain-specific differences in X and Y chromosome rDNA hybridization
          patterns detected using the Southern blotting technique. I n this report, I ex-
          tend the previous observations by examining polytene rDNA patterns in
          wild-type and hybrid X / Y cells. A dominance hierarchy for the X and Y
          chromosomes from three strains of D. melanogaster is presented and possible
          mechanisms of replicative dominance are discussed.

  N Drosophila melanogaster the genes than encode the 18s and 28s ribosomal
I RNAs are located at the nucleolus organizer regions on the X and Y chromo-
somes (RITOSSA   and SPIEGELMAN   1965). The number of genes at each nucleolus
organizer site has been estimated to be 200-250 (TARTOF    1971, 1973).
   Evidence from rRNA saturation hybridization experiments indicates that the
number of ribosomal gene repeats present in certain somatic tissues of Droso-
phila may differ from the germline number of genes. In particular, HENNIG
and MEER(1971) and SPEAR       and GALL(1973) have shown that the ribosomal
genes in polytene tissue are under-replicated with respect to the bulk of the
DNA sequences. SPEAR     and GALL(1973) further demonstrated that while X / O
diploid cells contain half as much rDNA as X / X diploid cells, the number of
ribosomal genes present in X / O polytene tissue is the same as in X / X polytene
tissue. They suggested that the ribosomal genes in X / O tissue undergo an ex-
tra round of DNA replication during polytenization to compensate for the
lower starting number of genes. This would result in a doubling of the ribo-
somal genes in X / O cells to make rDNA levels in X I 0 and X / X polytene tissue
the same.
   We have shown that the major ribosomal gene hybridization pattern detected
in D.melanogaster Oregon-R X / Y polytene cells or in X / X polytene cells from
hybrids between D. melanogaster strains Canton-S, Oregon-R and OK-1 cor-
Genetics 100: 375-385 March, 1982.
3 76                                      S . A. ENDOW

responds to only one of the two nucleolus organizers present in the starting
diploid cell (ENDOW GLOVER
                       and          1979; ENDOW    1980). These findings suggest
that genes from only one nucleolus organizer undergo replication during poly-
tenization of X / Y or X / X cells. They also explain the finding of SPEARand
GALL (1973) that X / O and X / X polytene cells contain the same amount of
rDNA by suggesting that genes from only one nucleolus organization undergo
polytenization in either X / O or X / X cells, and that replication occurs to the
same extent in both cell types.
   In this paper I extended the previous findings by examining DNA from
diploid and polytene tissue of X / Y wild-type and hybrid flies. Results of these
experiments, together with previously published results (ENDOW       1980), allow
the construction of a dominance hierarchy for the Canton-S, Oregon-R and
OK-1 X and Y chromosomes. During this study it was found that X / Y flies
which carry a Canton-S X chromosome and an Oregon-R or OK-1 Y chromo-
some exhibit polytene rDNA patterns which are characteristic of both the X
and Y chromosomes. This observation may be accounted for in one of two
ways: 1) Genes from both nucleolus organizers may be replicated in each cell,
or 2) genes from only one of the two nucleolus organizers present may undergo
replication in each cell with random selection of the nucleolus organizer to be
replicated. While data supporting either of these possibilities was not obtained,
the second possibility is consistent with observations reported previously
 (ENDOW   and GLOVER     1979) and in this paper which indicate that in other
wild-type X / Y cells, genes from predominantly one nucleolus organizer undergo
multiple rounds of DNA replication during polytenization.

                                 MATERIALS A N D METHODS

   Drosophila stocks: Stocks of Drosophila melunoguster Ore-R A, Canton-S 1-1 and OK-l
were obtained, restarted through a series of single pair matings and characterized as described
previously (ENDOW     and GLOVER   1979; ENDOW  1980).
   Interstrain hybrids were constructed by mating a single virgin female of one strain with
a male from another strain. F1 hybrid X / Y progeny were selected for analysis.
                                                          A
   X k / ? or X / O flies were constructed by mating an X X female wlth an Ore-R A, Canton-
                            A
S 1-1 or OK-I male. The X X chromosome used in these studies was the C(I)RM, su(w")wayz
                                  A   A                    A
chromosome maintained in an X X / X Y stock with the X Y chromosome, YSX.YL, In(l)EN, dl
49, Y S car f U y.YL. This stock was originally obtained from F. RITOSSA
                                                                       (University of Bari).
       A
F, X X / Y females or X / O males were selected for analysis.
       A
   xx-,,/y     female flies were constructed by mating C ( I ) D X , Y f / B s Y females with Can-
ton-S 1-1 or OK-l males. Yellow female larvae were selected. Sibling adult females were
yellow with forked bristles and round eyes. C ( I ) D X , y f/Yo' flies which contain a Y chrom-
osome from the Ore-R A strain were from a stock of C ( I ) D X , y f/Yor/w"bbl flies. This stock
was constructed by mating a w"bbl/w%bc/BSY female to an Ore-R A male and selecting a
up, round-eyed F, male. This wabbl/OP male was mated to a C ( I ) D X , y f/YOr F, female
obtained from a cross between an Ore-R A male and a C ( I ) D X , y f/BSY female. The result-
ing stock consists of C ( I ) D X , y f/YO' females and w"bbi/Yor males, is stable and shows
characteristic rDNA patterns for DNA from diploid tissue of individual males or females.
    D N A eztraction: DNA was extracted from pooled samples of 12-50 sets of larval brains
and imaginal discs or salivary gland pairs as described previously (ENDOW        1980). In all cases
                          POLYTENIZATION O F      x A N D Y rDNA                         377
except one, pooled tissue samples were from progeny of a single mating pair. In most cases,
DNA samples from two or more separate matings were prepared and analyzed. DNA from
diploid and polytene tissue of XOr/O flies was a pooled sample prepared from progeny of five
single pair matings. Tissue was dissected in Drosophila saline (ROBB1969) and lysed in
0.5-1.0 ml of 0.5% sodium lauryl sarcosine (Sarkosyl), 0.1 M EDTA, 0.05 M tris-HC1 (pH
7.8). 0.91 g of CsCl was added for each 1.0 ml of lysate and EtBr (IO mglml) was added to
200 Pglml. Centrifugation time was 15-18 hr at 42,000 rpm and 20" in the Sorvall TV-865
rotor. When DNA was being recovered from very small tissue samples, centrifugation time
was extended to 20-24 hr to ensure tight banding of the submicrogram quantities of DNA.
   In some experiments, DNA from the brain and imaginal discs or from the salivary glands
of individual larvae was prepared for analysis according to procedures described previously
           and
(ENDOW GLOVER           1979).
    Southern transfers: Restriction of DNA with Eco RI, fractionation on 0.8% agarose gels,
transfer to nitrocellulose filter paper and hybridization with 32P-labelled Drosophila rDNA
were carried out as described previously (ENDOW       and GLOVER    1979; ENDOW  1980) with the
following modifications: The Drosophila rDNA probe was recombinant DNA pDm rea51 #1,
which consists of a noninterrupted 11.5 kb repeat unit. This recombinant was derived from
pDm r-a51 (DAWID,      WELLAUER LONG
                                  and        1978) which was kindly provided by E. LONG.   The
original pDm r-a51 clone contained two RI fragments in addition to the pMB9 vector. The
larger R I fragment was approximately 11.5 kb long and hybridized with a 32P-rDNA probe
that consisted of a n equimolar mixture of recombinant DNAs ckDml03A, ckDmlO3C, and
ckDmlO3D (GLOVER HOGNESS
                       and           1977). The smaller RI fragment of 6.5 kb did not hybridize
with our 32P-rDNA probe. Since the nature of the smaller fragment was unknown, we re-
cloned the 11.5 kb fragment by ligation into the RI site of pACYC184 (CHANG          and COHEN
1978) followed by transformation of E . coli HBlOl using the CaCls method. Colonies that
were tetR and CAPS were selected and hybridized with 32P-rDNA according to the colony
hybridization method of GRUNSTEIN HOGNESS
                                       and           (1975). DNA was prepared from a hybridi-
zation positive colony, pDm rea51 # I , and tested by digestion with restriction enzymes Hind
I11 and Eco RI, and by hybridization with ckDml03A, ckDml03D and ckDml03C. According
to these criteria, pDm r.a51 # I contains a complete uninterrupted rDNA repeat with no
other DNA sequences. It was therefore used as the rDNA probe in the experiments described
here. In most experiments, the rDNA insert was purified before in vitro labelling by digestion
of the recombinant DNA with Eco RI followed by agarose gel electrophoresis and electroelu-
tion of the band corresponding to the 11.5 kb rDNA fragment.
    The rDNA probe was labelled in nitro with 32P by nick-translation (KELLYet al. 1970;
MANIATIS,    JEFFREY  and KLEID1975) to specific activities of 107-108 cpm/pg. High background
on some filters led us to change our hybridization conditions and filter wash conditions to
those suggested by R. KAUFMAN       (unpublished). Filters were presoaked for 4-6 hr at 65" in
6 x SSC, 0.5% SDS, 2 x Denhardt's solution (1 x Denhardt's solution = 0.02% polyvinyl-
pyrrolidone, 0.02% ficoll and 0.02% bovine serum albumin; DENHARDT          1966), 10mM sodium
pyrophosphate, and 0.125 M sodium phosphate, p H 7. Filters were then transferred to 25 ml
of 3 x SSC, 2 X Denhardt's solution, 0.5% SDS, p H 7 and allowed to soak for 2 h r a t 65".
Denatured 32P-rDNA was added to this second soaking solution and hybridization was allowed
to proceed for 15-20 hr a t 65". Filters were washed at 65" in two changes (15 min each) of
3 x SSC, 2 x Denhardt's solution, 0.1% SDS, 0.1% sodium pyrophosphate, p H 7.2, followed
by two changes (1 hr each) of 1 x SSC, 0.1% SDS, 0.1% sodium pyrophosphate, p H 7.2 and
one change (30 min) of 0.2 x SSC, 0.1% SDS, p H 7. Following a brief rinse in 0.2 X SSC
at room temperature, filters were dried and exposed to X-ray film (Kodak XR-5).
     Filters containing DNA from the diploid tissue of individual larvae or from single pairs
of salivary glands were prepared and hybridized as described, except that 10% sodium dextran
sulfate (Pharmacia) was added to the second soak (and hybridization) solution (WAHL,STERN
and STARK1979). In some experiments sonicated, denatured pBR322 DNA was added to a
concentration of 3.2 Pg/ml to the second soak and hybridization solution. Care was taken to
agitate the filters frequently during the first two post-hybridization washes to remove the
viscous hybridization solution. Subsequent washes were carried out as usual.
378                                    S. A. ENDOW

                                         RESULTS

   Polytenization of ribosomal genes i X/O, X%.,/Y and X/Y flies: To deter-
                                       n
mine whether the ribosomal genes in X / O and X / X polytene tissue show the
same or different hybridization patterns, rDNA patterns for diploid and polytene
tissue of X / O and X / X flies were compared. DNAs from X / X Canton-S, Ore-R
and OK-1 flies show the diploid and polytene patterns reported previously
 (ENDOW    1980) and no detectable differences were found when X / O and X / X
diploid and polytene tissue were compared. In subsequent experiments, DNA
from X / X flies was used as a source of the X chromosome rDNA patterns.
   Hybridization patterns from X^x.xo/Y flies that contain rDNA from only
the Y chromosome were examined to determine the Y chromosome diploid and
polytene rDNA patterns. The Y chromosome rDNA patterns were then com-
pared with hybridization patterns for X / Y and X / X tissue from the correspond-
ing strain to determine whether the X / Y polytene pattern was derived from
the X or the Y chromosome or both. The major ribosomal gene repeat in all
of the Y chromosomes examined was 11.5-12 kb in length (Figure 1). In
 general, the Y rDNA patterns are strikingly different from the X rDNA pat-
 terns of the corresponding strain (ENDOW     1980). When DNA from polytene
tissue of X^x-so/Y flies carrying either an Ore-R or OK-1 Y chromosome was




     FIGURE l.-Kbosomal genes in diploid and polytene tissue of wild-type M Y flies. Auto-
radiograph of Southern transfer after digestion of DNA with F m R I and hybridization of
filter with SZP-,DNA. DNA was extracted from pooled samples of larval brains and imaginal
discs (diploid tissue) or larval salivary glands (polytene tissue). The lanes contain diploid
(d) or polytene (p) DNA from OK-1,Ore-R or Canton-S X/Y flies as indicated. DNA from
 A
XX-,dY flies carrying an OK-1,Ore-R or Canton-S Y chromosome (indicated as Y O K , Y '     o
or YC* DNA) and DNA from Canton-S X/X flies (lanes labelled XC8) are shown for com-
parison. The positions of the 17 kb and 11.5 kb repeats are indicated. Repeats which are pres-
ent in X/Y polytene cells but not in the corresponding Y chromosome polytene pattern are
indicated with a 0.
                       POLYTENIZATION O F   x AND Y r D N A                  379
examined, rDNA patterns found were almost identical to the corresponding,
                                                               A
X / Y polytene pattern (Figure 1 ) . Small differences in the XX-,o/Y and X / Y
polytene patterns were present. One such difference was the presence of a
minor amount of 15-16 kb repeat in the OK-I X / Y polytene pattern that was
absent from the $X-NO/YoK polytene pattern (Figure 1). Since this repeat
was at the position of one of the major X chromosome rDNA repeats, it may
have arisen from a low amount of X chromosome rDNA replication. The major
rDNA pattern present in OK-I X / Y polytene tissue, however, was the same
                      A
as that present in XX_,,/Yox polytene tissue. This suggests that genes from
predominantly the Y chromosome nucleolus organizer undergo replication in
OK-I X / Y polytene tissue. A similar observation has been reported previously
for Ore-R X / Y polytene tissue (ENDOW GLOVER
                                           and           1979). The Ore-R X / Y
                                                      A
polytene pattern shows some differences from the XX-No/Yo' polytene pattern
 (Figure 1). In particular, two high molecular weight repeats of > I 7 kb in Ore-
                                                                        A
R X / Y polytene tissue are not present in the same amount in XX.,o/Yor
R X/Y polytene tissue are not present in the same amount in XX-m/Yor
polytene tissue. These high molecular weight repeats probably represent partiaI
digestion products. Although digests were monitored using an internal standard,
a low amount of partial digestion products might escape detection. In both Ore-
R and OK-I X / Y polytene tissue, genes from the Y nucleolus organizer are
preferentially replicated and few X chromosome repeats appear to undergo
multiple rounds of DNA replication. In addition, many rDNA repeats of <11.5
kb in length that are present in low amounts on the Ore-R and OK-I Y chromo-
                                    A

somes are underreplicated in both XX-,,/Y and X / Y polytene cells.
   Examination of DNA from Canton-S X / Y polytene tissue revealed an rDNA
pattern similar to the Canton-S X chromosome polytene pattern (Figure 1) .
The Canton-S X / Y and X / X polytene patterns differ from one another in that
the amount of the 11.5-12 kb repeat is increased in X / Y polytene tissue, in-
dicating that some replication of the Canton-S Y chromosome rDNA has prob-
ably occurred. Ribosomal genes from the Y chromosome are not replicated to the
same extent as X-chromosome repeats, however, as judged by minor repeats
                 A
present in the XX_,,/Y polytene pattern but absent from the X / Y polytene
pattern. The predominance of the X-chromosome rDNA pattern in Canton-S
X / Y polytene cells contrasts with Ore-R and OK-I X / Y polytene tissue, which
show a predominance of the Y-chromosome rDNA pattern. These observations
indicate that the Y chromosome nucleolus organizer is replicatively dominant in
Ore-R and OK-I X / Y polytene cells, but that the X nucleolus organizer is
replicatively dominant in Canton-S X / Y polytene cells.
   Polytenization of ribosomal genes in hybrid X/Y cells: X / Y hybrids were
constructed with Canton-S, Ore-R and OK-I flies, and polytene rDNA patterns
were examined. Hybrids that carry an Ore-R or OK-1 X chromosome and
a Y chromosome from any of the three strains showed a polytene pattern
characteristic of the Y chromosome (Figure 2 ) . In some cases minor bands
were detected that could be attributed to replication of X chromosome riboso-
380                                      S. A. ENDOW




   FIGURE     2.-Polytenization of ribosomal genes in X / Y hybrid flies. Autoradiograph of
Southern transfer after digestion of DNA with Fko RI and hybridi7stion of filter with
3zP-rDNA. X/Y hybrid flies were constructed with flies from the strains Canton-S, Ore-R and
OK-1. DNA was extracted from samples of larval brains and imaginal discs (diploid tissue)
or larval salivary glands (polytene tissue) pooled from progeny of single pair matings. The
figure shows DNA from diploid (d) or polytene (p) tissue of X O K / Y O r , X O r / Y O K , Xor/Ycs
and X O K / Y C s flies, as indicated. The arrows indicate the position of the 11.5 kb and 17 kb
repeats. Major repeats derived from the X chromosome nucleolus organizer are marked
with a 0.

mal gene repeats or to partial digestion products. In each case, however, the
major rDNA pattern was that of the Y chromosome, indicating that ribosomal
genes from predominantly the Y chromosome undergo multiple rounds of rep-
lication in polytene tissue of these hybrids.
   When flies containing a Canton-S X chromosome together with a Y chromo-
some from either the OK-1 or Ore-R strain were examined, DNA from polytene
tissue showed an rDNA pattern characteristic of both the Canton-S X and Ore-
R or OK-1 Y chromosomes (Figure 3). This conclusion was based on the rela-
tive amounts of 13-14 kb Canton-S repeat and 11.5 kb OK-1 repeat present in
the Xcs/YoKpattern and the presence of both the 12 kb Ore-R and 11.5 kb
Canton-S repeats in the Xcb/Yorpattern. Examination of DNA from salivary
gland pairs of hybrid individuals yielded similar results. The finding of both
the X and Y rDNA patterns in hybrid polytene tissue suggests either that genes
from both nucleolus organizers are replicated to the same extent in each poly-
tene cell, or that genes from only one nucleolus organizer are replicated in each
cell and that selection of the nucleolus organizer to be replicated is random.
                          A
   Ribosomal genes in XX/Y polytene cells: The domina2ce effect in cells con-
taining multiple nucleolus organizers was examined in X X / Y cells that contain
three nucleolus organizers. DNA from X%/Y polytene tissue carrying an Ore-R
                          POLYTENIZATION OF       x AND Y rDNA                            381




   FIGURE  3.-Polytenization of ribosomal genes in X C s / Y O K and X C S P flies. Autoradio-
graph of Southern transfer after digestion of DNA with Eco R I and hybridiiation of filter
with s*P-rDNA. X/Y hybrid flies were constructed by mating a Canton-S X / X female fly
with an OK-1or Ore-R X / Y male fly. DNA was extracted from samples of larval brains and
imaginal discs (diploid tissue) or larval salivary glands (polytene tissue) pooled from progeny
                          h              A
of single pair matings. XX-No/YOK, X X - N d Y O r and X C s / X C s rDNA patterns (labelled
YoK, Yo' and X C S , respectively) are shown for comiarison. The lanes contain DNA from
diploid (d) or polytene (p) tissue of X / Y , X/X or XXeNo/Y flies as indicated. The arrows
show the positions of the 11.5 kb and 17 kb repeats.

or Canton-S Y chromosome showed only the Y chromosome polytene pattern
                                                                    A

as judged by comparison with DNA from appropriate XX.n.o/Y polytene cells
(Figure 4). These results indicate that genes from only one nucleolus organizer
                              h

undergo replication in X X / Y polytene tissue, although three nucleolus organ-
                            h               A
izers are present. In both X X / Y r * and X X / Y o r flies, Y chromosome ribosomal
                                                                                           A

genes are replicated in polytene tissue, while ribosomal genes from the X X
chromosome undergo little or no replication.

                                        DISCUSSION

  Results of analysis of X / Y wild-type and hybrid flies are summarized in
Table 1 together with results of analysis of X / X hybrid flies (from ENDOW
1980). These data allow the construction of a dominance hierarchy such that
the Canton-S X chromosome is replicatively equivalent to the Ore-R or OK-1Y
chromosome, since neither of these chromosomes can be repressed by the
Canton-S X chromosome. The Canton-S X chromosome is replicatively domi-
nant to its own Y chromosome, which in turn is dominant to the Ore-R and OK-
382                                     .
                                       S A. ENDOW




                                                L.




                                                      n
   FIGURE
        4.-Polytenization      of ribosomal genes in X X / Y flies. Autoradiograph of Southern
                                             !
                                             R
tr2nsfer after digestion of DNA with Eco , and hybridization of filter with **P-rDNA.
X X / Y flies were constructed by mating an X X female with a Canton-S or Ore-R male fly.
DNA was extracted from samples of larval brains and imaginal discs (diploid tissue1 or larval
salivary glands (polytene tissue) pooled from progeny of single pair matings. X X - , d W
                A                                 h
(labelled YC*), X x - ~ o / Y o ' (labelled YO') and X X rDNA patterns are shownAfor co%pan-
sy.   The lanes contain DNA from diploid (d) or polytene (p) tissue of X X / Y , X X or
X X - N d Y flies, as indicated. The positions of the 11.5 kb and 17 kb repeats are marked
with arrows.

I chromosomes; the OK-I X chromosome is dominant to the Ore-R X chromo-
  X
some. These relationships may be expressed as follows:
                      X' ,YOK , O > YC"> XO" > X'
                       C       Y'               O
The observations that the Canton-S X chromosome is dominant to the Ore-R
and OK-I X chromosomes in X/X hybrids, and that the Ore-R and OK-I Y
chromosomes are replicatively dominant to the Ore-R and OK-1 X chromosomes
are consistent with this scheme. The finding that genes from both nucleolus
organizers undergo replication in cells which contain a Canton-S X chromosome
and an Ore-R or OK-1 Y chromosome is consistent with the finding that the
Canton-S X chromosome is a very dominant chromosome that can suppress
replication of its own Y chromosome in X/Y polytene cells. The Canton-S X
chromosome thus exhibits the strong dominance characteristic of the Ore-R and
OK-1 Y chromosomes, but does not appear more dominant than either of these
two Y chromosomes. Whatever the mechanism by which this dominance is
mediated, the Canton-S X chromosome appears not to be suppressible. It may
lack a receptor for a repressor-like activity or, alternatively, it may show
tight binding to an activator substance.
                              POLYTENIZATION OF      x AND Y rDNA                  383
                                              TABLE 1
                                                                       A
                    Polytenization of ribosomal genes in X/Y, X/X and XX/Y flies




 2. X / X flies*




 3. X>/Y    flies



  * Data from ENDOW(1980).

   The construction of a dominance hierarchy suggests that a locus that can
exist as a series of alleles may be involved in mediating the dominant effect.
The molecular basis of this dominant effect may be the production of a series
of repressor-like proteins that bind with different affinities to a receptor site
and result in a blocking of DNA replication. Alternatively, the existence of
allelic activator proteins with different binding affinities for an origin of DNA
replication may be involved in mediating the dominant effect. A third possibility
is that the number of activator protein binding sites or origins of DNA rep-
lication may determine the relative "strength" of dominance.
            A
   When X X / Y cells that contain three nucleolus organizers were examined,
                                                                             A
diploid tissue showed ribosomal gene repeats characteristic of both the XX and
                                         A
Y chromosome nucleolus organizers. X X / Y polytene tissue, however, showed
only the Y chromosome polytene pattern, indicating that genes from only one
nucleolus organizer are replicated in cells containing multiple nucleolus orga-
nizers and again illustrating the strong dominance of the Y chromosome in
suppressing polytenization of X-linked ribosomal genes. Replication of genes
from only one nucleolus organizer in polytene cells of flies that contain
multiple nucleolus organizers was predicted from work of SPEAR       (1974). SPEAR
showed that rDNA levels in polytene cells that contained one, two, three or
384                                     S.    .
                                             A ENDOW
four nucleolus organizers were the same, in contrast to diploid cells, which
showed the expected 2-fold, 3-fold, or 4-fold increase in the amount of rDNA,
parallelling the increase in the number of nucleolus organizers. SPEAR    suggested
that one possible interpretation of this observation was that only one of the
nucleolus organizers undergoes polytenization in cells which contain multiple
nucleolus organizers, while the remaining nucleolus organizers are replicatively
inactive. Earlier, MACGREGOR    ( 1973) suggested that replicative inactivation OC-
curred during polytenization of Drosophila cells; this was also proposed by HEN-
NIG and MEER(1971) to explain underreplication of rDNA in polytene cells of
D.hydei.
   Another type of nucleolar dominance that affects formation of the nucleo-
lus organizer constriction in interspecific hybrids of Drosophila has been
described previously by DURICA      and KRIDER (1977, 1978). This dominant ef-
fect probably differs from the nucleolar dominance described here, in that
it more likely affects rRNA transcription than rDNA replication.
    This study was supported by US Public Health Service Grant No. GM 25583 and National
Science Foundation Grant No. PCM-8110183. Stocks of Canton-S and OK-l flies were origin-
ally obtained through Dr. G. DOVER  from the University of Cambridge collection. Stocks of
 A    A    A

X X / X Y ; XX-No/BSY/In(l)scB, cu and w"bbl/BSY flies were obtained from Dr. F. RITOSSA,
University of Bari.
    I thank Dr. E. LONGfor a gift of pDm r.a51 and C. RICHARDS assistance in construct-
                                                                for
ing pDm r.a51 # 1 . Drs. J. BOYNTON N. GILLHAM
                                       and           provided helpful discussions and criti-
cism. I am especially indebted to Dr. R. KAUFMAN suggesting our current filter hybridiza-
                                                for
tion and wash conditions and to Dr. P. MODRICH a generous gift of crystalline-pure Eco
                                                for
RI. I also thank D. STEPHENS a careful typing of the manuscript.
                               for

                                    LITERATURE CITED

CHANG, C. Y. and S. N. CONEN,1978 Construction and characterization of amplifiable
        A.
   multicopy DNA cloning vehicles derived from the P I 5 8 cryptic miniplasmid. J. Bac-
   teriology 134: 1141-1156.
DAWID,I. B., P. K. WELLAUER E. 0. LONG,1978 Ribosomal DNA in Drosophila mela-
                               and
   nogaster I. Isolation and characterization of cloned fragments. J. Mol. Biol. 126: 749-768.
        D.
DENHARDT, T., 1966 A membrane-filter technique for the detection of complementary
   DNA. Biochem. Biophys. Res. Comm. 23: 641-646.
UURIC.4, D. S. and H. M. KRIDER,    1977 Studies on the ribosomal RNA cistrons in interspe-
    cific Drosophila hybrids. I. Nucleolar dominance. Develop. Biol. 59: 62-74. - 1978
                                                                                   ,
    Studies on the ribosomal RNA cistrons in interspecific Drosophila hybrids. 11. Hetero-
    chromatic regions mediating nucleolar dominance. Genetics 89: 37-64.
       S.
ENDOW, A., 1980 On ribosomal gene compensation in Drosophila. Cell 22: 149-155.
ENDOW, A. and D. M. GLOVER,
       S.                         1979 Differential replication of ribosomal gene repeats in
   polytene nuclei of Drosophila. Cell 17: 597-605.
GLOVER, M. and D. S. HOGNESS,
       D.                         1977 A novel arrangement of the 18s and 28s sequences
   in a repeating unit of Drosophila melanogaster rDNA. Cell 10: 167-176.
           M.
GRUNSTEIN, and D. S. HOGNESS,      1975 Colony hybridization: A method for the isolation
   of cloned DNAs that contain a specific gene. Proc. Natl. Acad. Sci. U.S.A. 72: 3961-3965.
                          POLYTENIZATION O F     x AND Y rDNA                           385
HENNIG, and B. MEER, 1971 Reduced polyteny of ribosomal RNA cistrons in giant
      W.
    chromosomes of Drosophila hydei. Nature New Biology 233 : 70-72.
KELLY,R. B., N. R. COZZARELLI, P. DEUTSCHER, R. LEHMAN A. KORNBERG,
                               M.                   I.              and               1970
   Enzymatic synthesis of deoxyribonucleic acid XXXII. Replication of duplex deoxyribo-
   nucleic acid by polymerase at a single strand break. J. Biol. Chem. 245: 3 9 4 5 .
       H. C.,
MACGREGOR, 1973 Amplification, polytenization and nucleolus organizers. Nature New
    Biol. 246: 81-82.
MANIATIS, A. JEFFREY D. G. KLEID,1975 Nucleotide sequence of the rightward oper-
           T.,              and
   ator of phage A. Proc. Natl. Acad. Sci. U.S.A. 72: 1184-1188.
RITOSSA: M. and S. SPIEGELMAN,
         F.                            1965 Localization of DNA complementary to ribosomal
   RNA in the nucleolus organizer region of Drosophila melanogaster. Genetics 53 :737-744.
ROBB,J. A., 1969 Maintenance of imaginal discs of Drosophila melanogaster in chemically
   defined media. J. Cell Biol. 41: 876485.
SPEAR, B., 1974 The genes for ribosomal RNA in diploid and polytene chromosomes of
       B.
   Drosophila melanogaster. Chromosoma 48: 159-179.
SPEAR, B. and J. G. GALL,1973 Independent control of ribosomal gene replication in poly-
       B.
   tene chromosomes of Drosophila melanogaster. Proc. Natl. Acad. Sci. U.S.A. 70: 1359-
    1363.
TARTOF, D., 1971 Increasing the multiplicity of ribosomal RNA genes in Drosophila
         K.
    melanogaster. Science 171 : 294-297. - 1973. Regulation of ribosomal RNA gene
                                                   ,
   multiplicity in Drosophila melanogaster. Genetics 73 : 57-71.
WAHL,G. M., M. STERNand G. R. STARK,            1979 Efficient transfer of large DNA fragments
    from agarose gels to diazobenzyloxymethyl-paper and rapid hybridization by using dex-
    tran sulfate. Proc. Natl. Acad. Sci. U.S.A. 76: 3683-3687.
                                                               Corresponding editor: G. LEFEVRE

								
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