ISSN 10227954, Russian Journal of Genetics, 2011, Vol. 47, No. 12, pp. 1479–1486. © Pleiades Publishing, Inc., 2011.
Original Russian Text © L.N. Porokhovnik, V.V. Viktorov, N.A. Egolina, T.G. Tsvetkova, N.A. Lyapunova , 2011, published in Genetika, 2011, Vol. 47, No.12, pp. 1666–1675.
Ribosomal genes (RGs) occur in many copies in
the human genome and are localized in the nucleolus
organizer regions (NORs) of five acrocentric chromo
some pairs (13, 14, 15, 21, and 22) to form clusters of
various sizes. The total RG copy number per diploid
cells varies from 250 to 670 among individual human
genomes , and only some copies, whose number
varies among individuals, are transcriptionally active
in somatic cells . A specific silver nitrate (Ag) stain
ing of metaphase chromosomes [3, 4] reveals the active
RG (AcRG) clusters [5, 6]. The size of the silver pre
cipitate (AgNOR) can be estimated visually and
expressed in arbitrary units from 0 to 3 (Fig. 1) [7, 8].
Each of the ten NORs may have any of the four
AgNOR size variants. The characteristic is stable for
each chromosome, is reproduced through cell divi
sions within one tissue (Fig. 1b), and is the same in dif
ferent tissues (lymphocytes, fibroblasts) of one indi
vidual [9, 10]. In addition, the AgNOR size variants of
each chromosome are inherited as Mendelian charac
ters [11, 13]. This provides additional evidence for the
fact that the AcRG cluster sizes on metaphase chro
mosomes are the same in all cells of a given individual.
Since all AcRGs are functionally equivalent, the sum
of the ten AgNOR sizes was used as a characteristic of
the individual AcRG genomic dosage [7, 8], which is
thereby considered to be an additive quantitative trait
that is determined by five polymorphic autosomal loci
(with four allelic forms for each locus). The AgNOR
size was found to correlate with the RG copy number
in the cluster in the majority of cases [7, 14]. One arbi
trary unit corresponds to 8
1 AcRG copies .
We have previously used cytometry to measure the
sizes (areas) of the silver precipitates over NORs of
metaphase chromosomes. The results showed a close
correlation between visual estimates and cytometric
AgNOR size measurements . At the same time, it
was found that an automated analysis fails to yield a
stable average AgNOR size unless the number of mea
surements is extremely great and stringent require
ments are imposed on the quality of metaphase plates.
Because of this, AgNOR morphometry is unsuitable
for a mass analysis of NORs on human chromosomes.
At the same time, the results demonstrate the suitabil
ity of visual estimates of the AgNOR sizes.
Cluster Size Polymorphism of Active Human Ribosomal Genes
and Simulation of the Conditions of Its Stability through Generations
L. N. Porokhovnik, V. V. Viktorov, N. A. Egolina, T. G. Tsvetkova, and N. A. Lyapunova
Medical Genetics Research Center, Russian Academy of Medical Sciences, Moscow, 115478 Russia
Received September 9, 2010
—Based on selective silver nitrate staining of active ribosomal gene (AcRG) clusters in nucleolus
organizer regions (NORs) of human metaphase chromosomes, a technique was developed earlier to estimate
the AcRG dosage in individual genomes as a sum of arbitrary units (0–3) ascribed to the silver precipitate
(AgNOR) on ten NORs. The AcRG dosage was considered to be an additive quantitative trait determined by
five polymorphic autosomal loci (with four allelic forms for each locus). A database was created to contain
the data on AcRG cluster variants for more than 1000 individual human genomes. In this study, the popula
tion frequencies of AcRG cluster variants were determined. The results agreed with the hypothesis that sta
bilizing selection acts at the zygotic and/or early embryogenetic stage to restrain the AcRG genomic dosage
(copy number) within a range from 14.9 to 23.7 arbitrary units (the cell is unviable when the trait is beyond
this range). The average zygotic losses due to selection were estimated at 9.1–9.9% for a real population. A
computer model where the AcRG dosage of a progeny results from a random combination of the AgNORs of
the five acrocentric chromosome pairs of the parents was developed and used to simulate the formation of a
certain AcRG genomic dosage through generations in a human panmictic population with nonoverlapping
generations. A combination of stabilizing selection by total AcRG copy number and a certain spontaneous
mutation rate (the probability of changes in the cluster size of a NOR as a result of unequal crossingover in
meiotic prophase) was shown to be a sufficient condition for the restrain of equilibrium population frequen
cies of AgNOR size variants in a human panmictic population. Using the model, the most probable sponta
neous mutation frequency was predicted to be (2.1–2.3)
per NOR per generation for human AgNORs.
The predicted frequency was within the 95% confidence interval of the experimental rate, which was deter
mined by studying the inheritance of AgNOR variants in real families.