Identifying genes that confer resistance to ultraviolet radiation

in the unicellular alga Chlamydomonas

Bryan LaBore

Northern Arizona University

Mentor: Dr. Karen VanWinkle-Swift

Why is this research important?

•Prevent damage caused by UV (UV-C & UV-B)

•Increasingly important with hole in ozone

•Damage to:•DNA mutations skin cancer•Protein defective enzymes altered metabolism•Lipids leaky membranes cell death

Harmful Effects of UV-B Radiation

• Avoidance (sunscreens)– Common mechanism in plants– Melanin in humans, and topical sunscreens

• Repair systems for DNA damage• Degrade and re-synthesize proteins and lipids

Protection from UV Radiation

Chlamydomonas as a model

• Unicellular green alga

• Photosynthetic• Requires sunlight

uvs-1 mutant

How we got the uvs-1 mutant

mutagen

WT uvs-1

0 sec

15 sec

30 sec

45 sec

Survival after exposure to UV radiation

0 25 50 750.001

0.01

0.1

1

10

100wildtype

uvs-1

UV dose (seconds)

Wild-type vs. uvs-1 Survival at Various Doses of UV Radiation

New sup mutation

How we got the sup mutant

Super sunscreen?Super repair?

mutagen

UVS supsvegetative cells

0 10 20 30 40 50 60 70 80 90 1000.0001

0.001

0.01

0.1

1

10

100u220

uvs

18-3

16-1

8-2

3-2

9-1

4-1

2-2

25-1

15-2UV dose

(continuous light protocol)

Suppressor mutants have regained resistance to UV radiation

UV Dose(sec)

Now What?• What does the suppressor mutation do? • We must isolate the sup mutation from the uvs mutation, which requires some basic genetics

A Review of Basic Genetics

Parentals Recombinants

Defective UV repair? (uvs)

Super sunscreen? (sup)

Genetic Analysis of the UV resistant suppressor mutants

Normal UV repair (not uvs)

Normal sunscreen (not sup)

Normal

NormalUV resistant (uvs sup)

UV resistant (uvs sup)

UV sensitive (uvs)

SUPER UV resistant?!?! (sup)

Further Research To Be Done

• Isolate the sup mutation in all of the sup mutants• Perform further survival curves on the isolated mutants•Determine the mechanism behind each sup mutation (Super sunscreen? Super repair?)

Thank you to:• NASA• NAU• Dr. Nadine Barlow• Kathleen Stigmon• Dr. Karen VanWinkle-Swift

Questions?