In Situ Localization of Gene-Specific Transcription Regulatory Factors by Southwestern Histochemistry (<Symposium XII>Hybridohistochemistry)
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To understand better the regulation of gene expression, an analysis of the expression of trans-acting transcription regulatory factors in individual cells is essential. Although cellular analysis of specific mRNA expression is successfully performed by in situ hybridization, the results are not enough to tell 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. Recently, we have developed a new method, Southwestern histochemistry, which allows us to localize transcription regulatory factors in situ. In principle, double-stranded (ds) DNA harboring specific response element base sequences is synthesized and labeled with a hapten such as thymine-thymine (T-T) dimer and digoxigenin (Dig). After the reaction of frozen tissue sections with the probe ds DNA, the signal is detected enzyme-immunohistochemically by horseradish peroxidase (HRP) -labeled anti-T-T dimer or anti-Dig, respectively. As a model system, we localized estrogen receptor (ER) in mouse uterine tissue sections by immunohistochemistry with 1D5 monoclonal antibody and Southwestern histochemistry using T-T dimerized estrogen responsive element (ERE) ds DNA of vitellogenin. Consequently, very similar localization of ER was obtained by both methods. This result, together with other examples, further indicates the usefulness of Southwestern histochemistry as a method to localize specific transcription regulatory factors in situ.
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