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The nucleolar organizer in the mitotic chromosome complement of Xenopus laevis
By J. KAHN
(From the Medical Research Council Radiobiological Research Unit,
Harwell, Didcot, Berkshire)
With one plate (fig. i)
Summary
The mutation which reduces nucleolar number in interphase nuclei of Xenopus laevis
also reduces the number of secondary constrictions in mitotic chromosome complements.
Introduction
T H E diploid interphase nuclei of wild-type Xenopus laevis each contain a
maximum number of 2 nucleoli. The 2 nucleoli can be seen in most cells as
two distinct bodies, but sometimes they are fused and form only one. Wickbom (1945) reported the diploid chromosome number of X. laevis to be 36.
In 1958 Elsdale, Fischberg, and Smith described a mutation that reduces
nucleolar number in X. laevis. They observed that half the offspring from
a particular diploid female never possessed more than a single nucleolus in
each diploid nucleus. Breeding tests showed that these i-nucleolate mutants
are heterozygotes which behave in a simple Mendelian manner. Homozygous
enucleolate larvae have been obtained, but these die at the stage when wildtype and heterozygous larvae begin to feed (Fischberg and Wallace, i960;
Wallace, i960).
McClintock (1934), working with plant material, and Beermann (i960),
working with animal material, have shown that nucleoli normally arise at
specific regions on specific chromosomes. These chromosome regions, known
as nucleolar organizers, can be seen at mitosis as secondary constrictions.
Mutants with a reduced number of nucleolar organizers were found both by
McClintock and by Beermann. With their observations in mind I have
compared the mitotic chromosomes of normal wild-type Xenopus larvae with
those of the 1 -nucleolate heterozygotes.
Material and methods
The material used for this study, kindly provided by Professor M. Fischberg, was obtained from a cross between a heterozygous female and a wildtype male. It thus consisted of approximately equal numbers of wild-type
and heterozygous larvae.
Aceto-orcein squash preparations were made from gut and heart taken from
[Quart. J. micr. Sci., Vol. 103, pt. 4, pp. 407-9, 1962.]
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Ff
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Kahn—Nucleolar organizer of Xenopus
larvae on the first day of feeding. Nucleoli were readily seen in these orceinstained preparations.
Cytological observations
The diploid complements of both wild-type and heterozygous Xenopus
consist of 36 chromosomes. The complement of wild-type animals includes
a pair of chromosomes, both of which possess a secondary constriction. In
heterozygous animals only one such chromosome is present. This difference
in the number of secondary constrictions between 1- and 2-nucleolate animals
was found to be consistent throughout the material investigated (more than
20 larvae of each kind); it is therefore reasonable to assume that the constrictions mark the sites of nucleolar organizers.
Fig. 1, A and c, show respectively an interphase nucleus and a mitotic
metaphase of a wild-type larva. The nucleus in fig. 1, A, contains two
nucleoli; in fig. 1, c, attention is drawn by arrows to a pair of chromosomes
with subterminal centromeres. Each of these chromosomes has a secondary
constriction in its short arm close to the centromere. Fig. 1, B and D, show
respectively an interphase nucleus with a single nucleolus and a mitotic
prometaphase of a heterozygous larva: only one chromosome bearing a
secondary constriction is present, and it is marked with an arrow.
Measurements made on one mitotic complement of a heterozygous larva
indicate that the 8 largest chromosomes (4 pairs) have median, or nearly
median, centromeres. The 4 next largest chromosomes have subterminal
centromeres, with a great disparity between the lengths of the longer and
shorter arms, and they include the one chromosome which bears a secondary
constriction. The rest of the complement has not been analysed in detail.
Discussion
One might have expected those Xenopus whose nuclei form only one nucleolus apiece either to be aneuploid, lacking one of the 2 nucleolar-organizing
chromosomes, or to be euploids in which the nucleolar-organizing region of
one chromosome is, for one reason or another, defective. This study shows
that the latter supposition is correct.
I am grateful to Professor M. Fischberg and to Dr. A. Blackler for the
generous supply of material. I should also like to thank Dr. C. E. Ford for
his advice on the preparation of the manuscript; and Mr. B. Reeves for
skilful technical assistance.
FIG. 1 (plate), A, interphase nucleus from the heart of a wild-type larva of X. laevis showing
two nucleoli.
B, interphase nucleus from the gut of a heterozygous mutant with only one nucleolus.
C, mitotic cell from the same preparation as A. The two chromosomes each bearing a secondary constriction are indicated by arrows.
D, mitotic cell from the same preparation as B. An arrow marks the one chromosome bearing
a secondary constriction.
Kahn—Nucleolar organizer of Xenopus
409
References
BEERMAN, W., i960. Chromosoma, n , 263.
ELSDALE, T. R., FISCHBERG, M., and SMITH, S., 1958.
Exp. Cell Res., 14, 642.
FiSCHBERG, M., and WALLACE, H., i960. 'A mutation which reduces nucleolar number in
Xenopus laevis', in The cell nucleus, edited by B. Holmes. London (Butterworth's Scientific Publications).
MCCLINTOCK, B., 1934. Z. Zellforsch., 21, Z94.
WALLACE, H., i960. J. Embryol. exp. Morph., 8, 405.
WICKBOM, T., 1945. Hereditas, Lund, 31, 241.