Chapter 17
Introduction to the Detection of Transcription
Regulatory Proteins
T AKEHIKO KOJI
Does an increase in the level of a certain mRNA mean the increase
in transcriptional activity?
We have already described how to localize specific mRNA and
DNA in cell or tissue preparations in the previous section. As in
the case of northern blot analysis of mRNA expression, it is apparent that the information on the expression of mRNA by in situ
hybridization in itself represents only a steady-state level of gene
expression. Consequently, we cannot know whether an elevated
level of certain mRNA is due to an increased rate of transcription
of the gene or due to a decreased rate of degradation of the
mRNA. Thus, for a more precise understanding of the regulation
of gene expression, one would require an analysis of the expression of trans-acting transcription regulatory factors at the individual cell level. Those proteins, such as c-Myc (Kerkhoff et al.
1991), c-Jun (Curran and Franza 1988), CREB (Meyer and Habener 1993) and various steroid hormone receptors (Beato 1989),
bind specific consensus sequences in nuclear DNA known as responsive element and regulate transcriptional activity of the
genes with the element. To date, however, in most of the transcription regulatory factors reported so far, there is only a little
information on their cellular distribution because of the limited
availability of specific antibodies which allow us to perform immunohistochemical demonstration of those proteins. Considering that the expression level of those proteins is generally too low
to be isolated at a sufficient amount to raise antibodies against
them, we needed to explore a new methodology.
Correspondence to Takehiko Koji, Nagasaki University, Dept. of Histology
and Cell Biology, 1-12-4, Sakamoto, Nagasaki, 852-8523, Japan (phone +81095-849-7025; fax +81-095-849-7028; e-mail tkoji@net.nagasaki-u.ac.jp)
Introduction to the Detection of Transcription
Regulatory Proteins
T AKEHIKO KOJI
Does an increase in the level of a certain mRNA mean the increase
in transcriptional activity?
We have already described how to localize specific mRNA and
DNA in cell or tissue preparations in the previous section. As in
the case of northern blot analysis of mRNA expression, it is apparent that the information on the expression of mRNA by in situ
hybridization in itself represents only a steady-state level of gene
expression. Consequently, we cannot know whether an elevated
level of certain mRNA is due to an increased rate of transcription
of the gene or due to a decreased rate of degradation of the
mRNA. Thus, for a more precise understanding of the regulation
of gene expression, one would require an analysis of the expression of trans-acting transcription regulatory factors at the individual cell level. Those proteins, such as c-Myc (Kerkhoff et al.
1991), c-Jun (Curran and Franza 1988), CREB (Meyer and Habener 1993) and various steroid hormone receptors (Beato 1989),
bind specific consensus sequences in nuclear DNA known as responsive element and regulate transcriptional activity of the
genes with the element. To date, however, in most of the transcription regulatory factors reported so far, there is only a little
information on their cellular distribution because of the limited
availability of specific antibodies which allow us to perform immunohistochemical demonstration of those proteins. Considering that the expression level of those proteins is generally too low
to be isolated at a sufficient amount to raise antibodies against
them, we needed to explore a new methodology.
Correspondence to Takehiko Koji, Nagasaki University, Dept. of Histology
and Cell Biology, 1-12-4, Sakamoto, Nagasaki, 852-8523, Japan (phone +81095-849-7025; fax +81-095-849-7028; e-mail tkoji@net.nagasaki-u.ac.jp)