WEBVTT

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Good afternoon, everyone.

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My name is Bea
Middleton, and I'm

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a senior and neuroscience
major here at Mount Holyoke

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Like Pho, I spent my
summer studying proteins.

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But instead of studying
them computationally

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with a computer, I was
working in a wet lab--

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so studying actual
proteins in living cells.

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I was a summer student fellow
at Woods Hole Oceanographic

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Institution, working
in the Hahn Lab,

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studying how animals
sense and adapt

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to the different chemicals
in their environment.

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And this ties in
with my title, "When

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Poisons Overtake Your Home--

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How the Atlantic
Killifish Adapted

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Resistance to the Chemicals
in its Environment.

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So what are some of these
chemicals or toxicants

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that I'm talking about?

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PCBs, or Polychlorinated
Biphenyls,

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are a toxic waste product
of the electrical industry,

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and they currently pollute New
Bedford Harbor, Massachusetts.

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Here, there's a
population of killifish

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that, over many generations,
have developed a resistance

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to these PCBs.

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And this is a resistance that
the fish of the clean reference

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site, Scorton Creek,
Massachusetts, do not have.

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So how is this possible?

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What makes the New Bedford
Harbor population resistant?

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This is our question.

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The Hahn Lab
believes it has a lot

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to do with the Aryl
Hydrocarbon Receptor,

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or AHR, because this protein
mediates the toxic response.

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So in other words, it binds
to the chemical, like a PCB,

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and, in its now activated form,
becomes a transcription factor.

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So it goes into the
nucleus, binds to the DNA,

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and alters gene
regulation, leading

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to toxic effects on the cell.

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But this still doesn't
really answer our question

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because if both the resistant
and the non-resistant fish

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have this protein, what makes
them respond to the chemicals

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differently?

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Well, it turns out there's a
lot of variation among AHRs.

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There are actually four
different AHR genes or types

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of AHRs.

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And for any given type,
there's a number of variants,

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structural variants.

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Interestingly, two
variants of AHR2a

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are found more frequently
among the resistant population

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of New Bedford Harbor.

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So that's the
orange and the blue

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there that I'm referring to.

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And another two, the light
pink and the light blue,

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are found more frequently
in the sensitive population

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of Scorton Creek.

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So this should made you wonder,
is this structural difference

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between the variants of the
two different populations

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underlying a
functional difference

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that could explain why
one population's resistant

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while the other
one is sensitive?

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And this ties in with my
job for the summer, which

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is to measure the activity
of the different AHR

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variants in response to
treatment with a given

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toxicant, like a PCB.

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To do this, well,
if you remember,

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the AHR is a
transcription factor.

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So if I want to
measure its activity,

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I'm going to measure the
amount of transcription that's

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occurring in the cells.

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To measure the amount
of transcription,

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I use a luciferase assay, which
essentially lets me use light

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as a proxy for AHR activity.

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I'm going to walk
you through some

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of the basic molecular
biology behind this technique

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just so you can appreciate just
how cool this technique is,

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and it'll also give
you a better picture

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of what I was doing on a
day-to-day basis in lab.

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So I start out with
a cell culture,

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and I transpect with the
AHR variant of interest,

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where I put into the cell that
protein that I'm interested in.

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And along with it, I
put in this piece of DNA

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so that when the
AHR is activated

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when I treat with the toxicant,
it actually binds to the DNA

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and turns on transcription
of this luciferase gene,

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producing luciferase
protein in the cell.