Differential display technology is a mechanism based strategy by which alterations in gene expression are identified in eukaryotic cells. Careful choice of an experimental system with well controlled samples is essential for identifying relevant genes whose expression might fit into a mechanism or hypothesis. Moreover, years of effort may still be required to unravel the biological functions of these genes. Recently, this hard work of characterization is beginning to shed light on the biological function of genes identified by Differential Display. Listed here are several exciting works published recently in high quality peer-reviewed journals.

GenHunter would like to congratulate these researchers for their great work that helps to validate the power of differential display technology. GenHunter is proud of the role our products have played in the successful use of this method. We wish them best of luck and continued success.

1. "Identification of a novel vertebrate circadian clock-regulated gene encoding the protein nocturnin". Carla B. Green and Joseph C. Besharse. Proc. Natl. Acad. Sci. USA. (1996) 93:14884-14888.
This work is one of the finest examples for the use of the differential display method to identify changes in gene expression, in this case, caused by an organism's day-night cycle which is also called the circadian clock. Using photoreceptors of frog retina where the circadian clock is located and set, Drs. Green and Besharse from University of Kansas Medical Center compared the pattern of gene expression in the retina under complete darkness post-synchronization by day-night cycles. They identified a putative transcription factor which they call nocturnin (stands for night factor) whose expression is turned on immediately following the night cycle. They showed that nocturnin gene expression is regulated by a transcriptional mechanism and its expression in the animal is restricted to the retina. The rhythmic expression and the nature of nocturnin suggest that this gene may be a major component or effector of the circadian clock.

Further readings of interest:
"Use of a high stringency differential display screening for identification of retina mRNAs that are regulated by a circadian clock". Carla B. Green and Joseph C. Besharse. Mol. Brain Res. (1996) 37:157-165.
"Circadian timekeeping: Loops and layers of transcriptional control" Charles J. Weiz. Proc. Natl. Acad. Sci. USA. (1996) 93:14308-14309.

2. "Wild-type p53 negatively regulates the expression of a microtubule-associated protein". Maureen Murphy, Adrian Hinman and Arnold J. Levine. Genes & Development. (1996) 10:2971-2980.
Much of the work on the p53 tumor suppressor gene has been focused on its transcriptional activation of the down stream genes, with p21 cyclin dependent kinase cloned by subtractive hybridization being the best example. However, none of the genes identified so far as p53 target genes can account for the tumor suppression activity of p53. Using differential display, Dr. Arnold Levine, one of the discovers of p53, and his colleagues from Princeton University attempted to identify genes whose expression may be down regulated instead of activated by the wild-type p53 tumor-suppressor protein. In this article they report the identification of a gene called MAP4, a protein previously associated with microtubules. They showed that agents which can inhibit the p53-mediated transcriptional repression and apoptosis also can abolish the alteration in MAP4 expression by wild-type p53. By ectopic expression of MAP4 in cells that are induced to undergo apoptosis, they were able to show that MAP4 expression could significantly delay this process, suggesting that the negative regulation of MAP4 by p53 may be necessary for rapid progression of cell death.

3. "Secretory leukocyte protease inhibitor: a macrophage product induced by and antagonistic to bacterial lipopolysaccharide". Fen-yu Jin, Carl Nathan, Danuta Radzioch and Aihao Ding. Cell. (1997) 88:417-426.
Septic shock is an often lethal response to endotoxin or bacterial lipopolysaccharide (LPS) as a result of bacterial infection. The molecular mechanism that mediates this response which causes tens of thousands of deaths every year is still obscure. Dr. Ding and colleagues from Cornell University Medical College reported in this paper the identification of a secretory leukocyte protease inhibitor (SLPI), a key molecule which may be responsible for the individual difference in the degree of responsiveness to LPS. Since macrophages are the most responsive cell type to LPS stimulation, Dr. Ding's group used differential display to compare the pattern of mRNA expression in macrophage cell lines derived from two strains of mice congenic for a locus affecting LPS sensitivity. They successfully identified a gene previously identified as a secretory leukocyte protease inhibitor which is expressed constitutively in LPS insensitive mice but absent in LPS responsive mice. They showed that ectopic expression of SLPI in LPS responsive cells could suppress their LPS responsiveness, as measured by the activation of NF-kB and production of nitric oxide and TNFa. It is known that interferon-g could restore the LPS responsiveness in macrophages from LPS hyporesponsive mice. This work demonstrates that the effect of interferon-g is mediated by the down regulation of SLPI expression, thus rendering the cells sensitive to LPS.