Keywords: human heterochromatin
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Inside the nucleus of a mammalian cell preparing for cell division, the chromatin strands are long, thin, fragile, and twisted helices that curve, loop, and tangle with each other — like noodles in a bowl of soup. Given this analogy, how can the cell separate the noodles into two identical portions, without breaking any? In eukaryotic cells, this problem occurs during every mitotic phase a cell enters. To resolve the mess, DNA must be condensed into discrete, compact form: chromosomes.
Robin Allshire's laboratory developed a robust and clever genetic assay to identify proteins involved in the heterochromatin formation at S. pombe centromeres. In this experimental system, they inserted a marker gene called ade6 + into a pericentromeric territory near the central domain, within outer repeats (Figure 1b). This region of the fission yeast genome typically exhibits heterochromatin organization in wild-type cells. Any genes from this region should not be expressed if heterochromatin formation is maintained.
How did these researchers monitor the expression of ade6 + inserted in the centromeric region? There is a simple assay system for assessing genetic activity in fission yeast. If the reporter gene remains silent, the cells are defective in synthesizing adenine. When grown in media without sufficient supply of adenine, red-colored yeast colonies appear, due to accumulation of red intermediates in the metabolic pathway. Normally, yeast colonies would be white-colored if the gene was active. So a color comparison can show at a single glance whether or not the gene in question is active in the yeast colonies. Interestingly, partial alleviation of the silencing state (meaning partial expression) can result in intermediately-colored pink colonies (Figure 4). Duggan and Tang used this system to monitor gene silencing.