The source of data in the present work was UniGene,
virtually containing all the ESTs sequenced so far from a number of
different human tissues and pertaining to more than 90,000 human genes.
The correspondence between UniGene entries and genes is generally
assumed.
The quality of the data analyzed and produced by the present study is
partially dependent on the quality of UniGene data. Although UniGene is
the less redundant among the gene indexing databases,
15 some UniGene clusters can include sequences of chimeric clones
belonging to different genes, and very large genes can be represented
in UniGene by two or more clusters. For this reason, before
constructing the genomic map of transcripts, we decided to discard all
the UniGene clusters showing multiple chromosomal locations.
The complementary situation (different UniGene clusters for a unique
human gene) is also possible, especially when dealing with different
UniGene clusters showing no similarity with any known sequence and
mapped to the same position. However, this situation is expected to be
rather rare.
The total number of genes expressed in retina is probably between
10,000 and 30,000.
2 Presumably, fewer than 70,000 genes
are active in a differentiated tissue. Therefore, the sample of 4974
individual transcripts considered by the present study corresponds, at
worst, to approximately 7% of the presumed total number of genes
expressed in retina, an adequate size for statistical inference.
The computational approach to the analysis of transcriptional profiles
is based on the assumption that the level of activity of a given gene
may be inferred from the information regarding the number of the
corresponding ESTs. The impossibility to detect differences in gene
expression resulting from posttranscriptional regulatory processes is a
strong limitation of this approach, but the same bias is shared by all
the present methods for estimating individual gene expression on a
large scale.
EST sequencing, from which the in silico approach is derived, is
intrinsically inadequate to identify truly rare genes. However, when
the sample size is sufficiently large, a fairly good quantitative
estimation of the transcription level of highly or moderately expressed
genes is possible.
16 On the contrary, quantitative
hybridization on high-density cDNA array fails to detect 80% of the
transcripts identified in a given tissue by EST
sequencing.
17
The transcriptional profile of the retina is characterized by a low
number of highly expressed genes, accounting for less than 10% of the
total number of genes, but for approximately 50% of the detected
transcriptional activity. The percentage of tissue-specific genes
(9.8%) is higher than previously reported in human adipose
tissue
10 and five times higher than observed in skeletal
muscle.
6 The difference could be ascribed to the high
specialization of the retinal tissue and/or to the relative absence of
contaminant tissues in the sample from which the cDNA libraries were
prepared. The subsample including highly expressed genes and genes
found exclusively in retina could be profitably used for monitoring
major changes in the transcriptional profile, after physiological or
pathologic modifications.
Data on the transcriptional profile of the human retina have been
reported in two independent studies. However, the small size of the
sample reported in BodyMap
3 and the different type of
source cDNA library
4 make it impossible to perform a
statistical comparison with the present data.
More than 60% of genes included in the catalog showed a precise and
unique map assignment. The distribution of genes by chromosome
significantly differed from the expectation. In a preliminary study, we
observed a peculiar concentration of retinal genes on chromosome
17.
18 This finding is confirmed by the present
investigation, based on a much larger number of map entries, which
suggests also a concentration of retinal genes on chromosome 19. The
short arm of chromosome 17 is known to be a hot spot to which several
phenotypically distinct retinal disorders have been
mapped.
19 The most recent version of the Human Gene Map
(HGM)
13 show that chromosome 17 is richer in genes than
expected under the hypothesis of a constant gene density along the
human genome. In the present study the expected distribution of genes
by chromosome was calculated according to HGM data. Therefore, the
hypothesis of a particularly high concentration of retinal genes on
chromosomes 17 and 19 is even stronger. A selective concentration of
genes in specific chromosomes was observed also in human skeletal
muscle.
11 18
Chromosomal maps showing the location of genes expressed in retina
represent a novel and important resource for positional cloning of
genes involved in retinal disorders. Most mapping information obtained
from UniGene resulted from radiation hybrids (RH) mapping,
which is, at present, the most precise method of gene mapping, if we
exclude the direct localization on the genomic DNA sequence. The
presence of several genes mapped to the same megabase distance from the
p-telomere probably reflects the actual concentration of genes in short
chromosomal segments. However, the width of the interval corresponding
to a single point of the present map is variable, because the physical
distance between the two flanking markers may be different. In spite of
this, the present genomic map of retinal genes is, at the moment, the
most precise representation of their cluster formations along each
human chromosome. Moreover, the resolution of the map is adequate to
detect which group of genes corresponds to a given interval to which a
human disease was mapped by linkage studies.
The in silico reconstruction of the transcriptional profile of the
adult human retina and the building of the genomic map of genes
expressed in this tissue provides for the first time a resource in
which functional and structural genomics of retina are integrated on a
large scale. It is hoped that this resource will speed up the
identification of genes involved in retinal disorders and will enable
innovative approaches to the study of retinal development, physiology,
and diseases.