Chemical Senses Vol. 30 No. suppl 1 © Oxford University
Press 2005; all rights reserved
Negative Feedback Regulation Ensures the One Neuron–One Receptor Rule in the Mouse Olfactory System
Department of Biophysics and Biochemistry, Graduate School of Science, The CREST and PRESTO Programs of Japan Science and Technology Agency, The University of Tokyo, Tokyo 113-0032, Japan
Correspondence to be sent to: Hitoshi Sakano, e-mail: sakano{at}mail.ecc.u-tokyo.ac.jp
Key words: allelic exclusion, feedback regulation, locus control region, mutually exclusive expression, odorant receptor genes
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Introduction |
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 Top Introduction Positive regulation of the...Negative regulation of the...ConclusionsReferences |
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In the mouse olfactory system, there are ~1500 odorant receptor (OR) genes clustered at ~50 different loci which are scattered among most of the chromosomes (Buck and Axel, 1991
;
Zhang and Firestein, 2002;
Godfrey et al., 2004). Like
the antigen receptor genes in lymphocytes, the mammalian OR genes are expressed in a
mutually exclusive (Malnic et al.,
1999;
Serizawa et al., 2000) and
monoallelic (Chess et al.,
1994;
Ishii et al., 2001) manner in
olfactory sensory neurons (OSNs). DNA rearrangement has long been thought of as a
possible mechanism for the allelic exclusion of the OR genes. However, mice cloned with
OSN nuclei excluded the possibility of irreversible gene translocation as a mechanism to
activate a single OR gene in each OSN (Eggan
et al., 2004;
Li et al., 2004). How is it,
then, that allelic exclusion is achieved in the olfactory system? Since OSNs expressing a
given OR gene converge their axons to a specific set of projection sites (glomeruli) on
the olfactory bulb (OB) (Ressler et al.,
1994;
Vassar et al., 1994;
Mombaerts et al., 1996;
Feinstein and Mombaerts, 2004),
odorous stimuli received in the olfactory epithelium (OE) are converted to a topographic
map of activated glomeruli on the OB (Mori et
al., 1999). Thus, the one neuron–one receptor rule forms the
genetic basis for OR-instructed axonal projection of OSNs. Here, we discuss possible
mechanisms that regulate single OR gene expression in each OSN in mouse. |
Positive regulation of the OR gene expression |
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TopIntroductionPositive regulation of the...Negative regulation of the...ConclusionsReferences |
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Three activation mechanisms have been considered for the choice of one member of the multigene family: DNA recombination, which brings a promoter and the enhancer region into close proximity; gene conversion, which transfers a copy of the gene into the expression cassette; and LCR, which interacts with only one promoter site. Irreversible DNA changes, i.e. recombination and gene conversion, have been attractive explanations for the single OR gene expression, because many parallels can be found between the immune and olfactory systems. We have analyzed the nuclei of OSNs expressing the MOR28 by fluorescent in situ hybridization (FISH) (Ishii et al., 2001
). Both DNA and RNA FISHs were performed to detect the
chromosomal location of the genomic sequence and the transcriptionally active site,
respectively. In our experiments, DNA FISH detected two genomic loci in the nuclei for
the MOR28, one for the maternal and the other for the paternal allele. RNA FISH
detected the MOR28 primary transcripts at only one of the two genomic sites.
These experiments not only confirmed the mono-allelic expression of OR, but also excluded
a possibility of gene conversion. If it were gene conversion, there should be three DNA
FISH sites: no additional genomic site was detected for the transcriptionally active
site. Of course DNA changes in nearby regions could not be ruled out in this experiment.
More recently, two groups have independently cloned mice with postmitotic OSN nuclei, and
found that the cloned mice generated normal repertoire of OSNs and showed no irreversible
DNA changes in the OR genes.
Since our FISH analysis did not support the gene translocation models, we searched
for the cis-acting DNA region that may regulate the single OR gene expression.
Using transgenic constructs in yeast artificial chromosomes (YACs), we have studied the
OR gene cluster containing the MOR28 gene (Serizawa et al., 2000). Sequence comparison of the
mouse and human genomes revealed a 2-kb homology (H) region 75kb upstream of the murine
MOR28 gene (Serizawa et al.,
2003). It was found that the shorter YAC constructs lacking the H region do
not express the transgenes. Attachment of the H region DNA to the upstream end restored
the expression of all transgenes in the cluster. These results indicated that the H
region is a cis-acting locus control region (LCR) that activates the MOR28
cluster.
It is important to ask how the expression of one particular OR gene within the
activated cluster is ensured. We propose that the activation complex formed in the LCR
interacts with only one promoter site in the OR gene cluster. Such a mechanism has been
reported for the mutually exclusive expression of the human visual pigment genes in cone
cells (Wang et al., 1999;
Smallwood et al., 2002).
|
Negative regulation of the OR gene expression |
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TopIntroductionPositive regulation of the...Negative regulation of the...ConclusionsReferences |
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To avoid concurrent expression, the activation processes for the OR gene expression may be relatively slow and rate limiting. However, this would not preclude the possibility of a second OR gene activation in other OR gene clusters. To achieve the mutually-exclusive expression, we assumed that a negative feedback regulation is taking place in OSNs like in the immune system (Martensson et al., 2002
). To test this hypothesis,
we deleted a coding-sequence of the MOR28 transgene (del-MOR28), and
examined whether mutually exclusive expression is violated in the OSNs expressing the
del-MOR28 (Serizawa et al.,
2003). We found that the del-MOR28 transgene permitted the
activation of other OR genes in the transgene-expressing OSNs. Similar observations were
also made with frame-shift mutants, in which nucleotide deletions or additions in the
coding regions caused the frame shift and created stop codons in the downstream region.
These results suggest that the OR gene product—not mRNA but most likely
protein—has a regulatory role in preventing the secondary activation of other OR
genes. |
Conclusions |
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TopIntroductionPositive regulation of the...Negative regulation of the...ConclusionsReferences |
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The mammalian OR genes are expressed in a mutually exclusive and monoallelic manner in OSNs. Such unique expression forms the genetic basis for the OR-instructed axonal projection of OSNs to the OB. We have identified the LCR for the mouse OR gene cluster containing MOR28 (Serizawa et al., 2003
). We assume that physical interaction between the LCR and
promoter ensures the expression of only one OR gene within the cluster. This model is
also attractive in that it reduces the likelihood of a simultaneous activation of two
different OR genes, from a probability among ~1200 genes to that among ~50 loci. Recent
transgenic experiments demonstrated an inhibitory role of the OR protein in preventing
further activation of other OR genes (Serizawa
et al., 2003;
Lewcock and Reed, 2004;
Shykind et al., 2004).
Stochastic activation of an OR gene by LCR and negative feedback regulation by the OR
gene product probably ensure the one neuron – one receptor rule in the mammalian
olfactory system.
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References |
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TopIntroductionPositive regulation of the...Negative regulation of the...ConclusionsReferences |
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