The Mechanics Of Gene Transcription

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The Mechanics Of Gene Transcription

http://www.medicalnewstoday.com/articles/93424.php

The molecular machinery behind gene transcription -- the intricate

transfer of information from a segment of DNA to a corresponding

strand of messenger RNA -- isn't stationed in special " transcription

factories " within a cell nucleus, according to Cornell researchers.

Instead, the enzyme RNA polymerase II (Pol II) and other key

molecules can assemble at the site of an activated gene, regardless

of the gene's position.

The findings, published in the journal Molecular Cell, are the

result of an ongoing collaboration between the laboratories of

T. Lis, the Barbara McClintock Professor of Molecular Biology and

Genetics, and Watt W. Webb, professor of applied physics and the

S.B. Eckert Professor in Engineering. Jie Yao, the paper's lead

author, recently finished his Ph.D. at Cornell under Webb.

Using multiphoton microscopy, a technique developed by Webb that

allows high-precision 3D imaging in living cells, the researchers

observed polytene chromosomes -- giant, multistranded chromosomes in

the salivary gland tissue of fruit flies that have hundreds of sets

of the genome instead of the usual two sets in conventional cells.

They activated heat shock genes, which protect cells from sudden

rises in temperature, and watched them in real time as they began to

be transcribed. The researchers also tagged Pol II with a

fluorescent marker to track its movements within the nucleus.

While some reports have suggested that activated genes move to a

specific nuclear location for transcription, the Cornell research

supports the traditional view that gene activation is not dependent

on movement to special locations, or so-called " transcription

factories, " said Lis.

" You see the genes decondense and fill up with polymerase, but

they're not moving anywhere -- they don't collect in a single

place, " he said. Instead, the transcription machinery assembles at

the called-upon locus, regardless of its position in the nucleus.

To test the generality of the findings beyond polytene nuclei to

common (but much smaller and more difficult to test) diploid cells,

the researchers used a technique called fluorescence in situ

hybridization, which allowed them to detect the location of specific

DNA sequences along a chromosome in fixed cells.

Looking at the location of co-regulated heat shock genes (genes that

are transcribed simultaneously), they found that co-regulated pairs

that occupied distinct sites before heat shock were no closer

together after heat shock. As in the polytene chromosomes, the genes

did not move to a single site for transcription.

And using fluorescence recovery after photobleaching -- another

method engineered by Webb -- the researchers found that over time

Pol II began to recycle itself within newly formed " compartments "

around the activated gene.

" At some point you accumulate enough polymerase that it feeds back,

so in a sense you've created a factory de novo " said Lis. " This is,

to our knowledge, the first demonstration of Pol II recycling at a

specific gene in vivo. "

Lis and colleagues are now looking at other molecules involved in

transcription to see if they behave similarly. " We're hoping to

develop new ways to really see, in vivo, how gene regulation works

mechanistically, " he said.