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Postreplication repair in Neurospora crassa.

R E Calza, A L Schroeder

Molecular & General Genetics : MGG

|January 1, 1982

Summary

This study on UV-irradiated Neurospora DNA repair mutants shows that while DNA synthesis is initially blocked, high molecular weight DNA is eventually restored. Pyrimidine dimers are identified as the cause of DNA synthesis inhibition and cell death.

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Area of Science:

Molecular Biology
Genetics
Biochemistry

Background:

Ultraviolet (UV) radiation induces DNA damage, primarily pyrimidine cyclobutane dimers.
DNA repair mechanisms are crucial for cell survival after UV exposure.
The Neurospora uvs-2 mutant is defective in excision repair, making it a model for studying postreplication repair.

Purpose of the Study:

To investigate the impact of UV irradiation on nascent DNA synthesis in the Neurospora uvs-2 mutant.
To elucidate the role of pyrimidine dimers in UV-induced DNA synthesis inhibition and cell lethality.
To explore potential postreplication DNA repair pathways in Neurospora.

Main Methods:

Isotopic pulse labeling of nascent DNA.
Alkaline gradient centrifugation to assess DNA molecular weight.
Alkaline filter elution to measure DNA strand breaks and repair.
Photoreactivation to reverse pyrimidine dimers.
Caffeine inhibition studies to probe repair pathways.

Main Results:

UV irradiation dose-dependently reduced nascent DNA size and labeling rate in uvs-2.
The mutant recovered synthesis of high molecular weight DNA 3 hours post-UV, despite depressed rates.
Photoreactivation mitigated DNA synthesis depression and UV-induced cell killing, confirming pyrimidine dimers as the cause.
Caffeine studies suggested partial inhibition of a single repair pathway or alternative postreplication repair routes.

Conclusions:

The Neurospora uvs-2 mutant exhibits a temporary block in DNA elongation/ligation following UV damage.
Pyrimidine cyclobutane dimers are the primary cause of UV-induced DNA synthesis defects and cell killing.
Evidence suggests the presence of alternative or compensatory DNA repair pathways in Neurospora, potentially modulated by caffeine.

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