The 'evolution' of a PhD project—from jumping genes to biochemical pathways

Traffic, Jan, 2007 by Henry Chung

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'WHAT'S THAT DOT?'

I would have happily continued my research into jumping genes and the gene regulation of Cyp6g1, if not for one of the control experiments that we performed. Control experiments are very important in scientific research as they are needed to confirm that the result from the actual experiment is due to the original hypothesis rather than some alternative, perhaps trivial explanation, or an error in the design of the experiment.

Evolution comes into the spotlight again. As our ancestors--from the dawn of life as simple single cell organisms--slowly evolved into the diverse forms of life that we see today, the landscape of our genome has changed dramatically. Some new genes are born and some of the old genes have disappeared. One way a new gene can arise is by a process called gene duplication. Gene duplication is a process where a single gene duplicates itself by a variety of mechanisms, giving birth to another gene with a similar DNA sequence. As time goes by, each of these two genes may share the same function or one of the pair might acquire a different function from the original. While they could be expressed in different tissues, their DNA sequence will still be very similar. That is the likely explanation for how a big gene family like cytochrome P450s arose: from a single P450 gene in an ancient single cell organism, it has undergone many duplication events since. The human genome contains around fifty-five P450 genes and the fly genome has around eighty-five. This process can be persistent and independent down many lineages in the tree of life.

I was about to conclude some the experiments for the Accord project when Phil Daborn asked if I had done any control experiments for the in situ hybridisation of Cyp6g1. I replied, 'What control?' 'A control to see if your Cyp6g1 RNA probe cross-hybridises to its closest paralogue, Cyp6g2,' he said to me. A paralogue is the gene that has the closest DNA sequence to another gene, probably because the two arise via gene duplication. Cyp6g2 is the closest paralogue to Cyp6g1 and they probably derive from one ancestral gene millions of years ago. If you remember how in situ hybridisation works, the design of the probe is based on the DNA sequence of the gene. As Cyp6g2 has a highly similar DNA sequence to Cyp6g1, there is a probability that probes designed for Cyp6g1 may pick up Cyp6g2 expression and vice versa. The control for this is doing the same in situ hybridisation experiments using a Cyp6g2 probe to see if the expression of Cyp6g2 is similar to Cyp6g1.

And so I did. After several attempts, I did not see any staining in the midgut, Malpighian tubules and fat body where Cyp6g1 is expressed. However, I did see a tiny blue dot near the brain of the larvae. 'Hmmm, that's interesting,' I thought, and I wondered what it was. Being relatively untrained in the anatomy of the larvae (I hesitated and decided not to use the term 'anatomically challenged'), I consulted Phil Daborn who identified the staining as part of the ring gland complex. Tamar Sztal, a fellow PhD student in the lab, who is probably the most proficient among the three of us in the anatomy of the larvae, remarked, 'That's the corpus allatum, the organ that produces juvenile hormone, one of the most important hormones in insect development!'