C8494 - Genome Instability

Learning outcomes

Upon completion of the course, students will understand: i) the genetic and molecular mechanisms of DNA damage and repair, checkpoint activation, and genomic instability; ii) the functions of disease-relevant protein factors; iii) mutations and molecular defects that lead to the development of human diseases such as cancer, neurodegeneration, aging, and immunological disorders. Furthermore, students will be able to conduct experiments to assess genomic instability using various methodologies.

Course contents

DNA damage and mutations. Damage to bases and to the sugar-phosphate backbone. Mechanisms of DNA damage: oxidizing, alkylating, and ionizing agents. Chemical agents and antitumor drugs.

Mechanisms of base damage repair. Photorepair, nucleotide excision repair (NER), base excision repair (BER), mismatch repair. Factors involved in repair mechanisms and related hereditary syndromes.

Non-B DNA structures as an endogenous source of genomic instability. G-quadruplex, R-loop, cruciforms, Z-DNA, triplex/H-DNA: formation, genomic distribution (telomeric regions, promoters, fragile sites), physiological roles (transcriptional regulation, recombination), and effects on replication fork progression.

Conflicts between transcription and replication (transcription-replication conflicts, TRC). Collisions between transcriptional and replicative machineries. Role of R-loops and resolution factors (senataxin, RNase H1/H2, topoisomerase I, helicases). Consequences: fork stalling/collapse, double-strand breaks, chromosomal instability.

Mechanisms of DNA break repair: homologous recombination and non-homologous end-joining. Factors involved in repair mechanisms and related hereditary syndromes.

The Fanconi Anemia (FA) pathway and interstrand crosslink repair. FANCD2-FANCI ubiquitination. Coordination with homologous recombination and with replication fork protection factors (e.g., BRCA1/2, RAD51).

Checkpoints activated by DNA damage. Kinase cascade: ATM/ATR and the role of p53. Checkpoint activation in response to fork stalling, TRC, and unresolved crosslinks.

Mutations in DNA repair genes and neurodegenerative diseases and cancer.

Laboratory: execution of experimental protocols to measure genomic instability and gene expression via fluorescence microscopy, real-time PCR, and interpretation of results. (Optionally: a demonstrative module on R-loop or G-quadruplex detection with specific antibodies could be considered, if compatible with course resources.)

 

Students with learning disorders and\or temporary or permanent disabilities: please, contact the office responsible (https://site.unibo.it/studenti-con-disabilita-e-dsa/en/for-students ) as soon as possible so that they can propose acceptable adjustments. The request for adaptation must be submitted in advance (15 days before the exam date) to the lecturer, who will assess the appropriateness of the adjustments, taking into account the teaching objectives.

Readings/Bibliography

Text book:

1 Capranico G, Martegani E, Musci G, Raugei G, Russo T, Zambrano N, Zappavigna V. Biologia molecolare, EdiSES (last edition).

2 RF Weaver, Molecular Biology, McGraw - Hill (last edition)

Moreover, I will distribute original scientific articles and reviews on the studied topic during the class.

At the start of the laboratory, a written protocol will be distributed for the execution of experimental activity.

Teaching methods

The course consists of lectures by the teacher and seminar of the students on specific topics (flipped class). The teacher will stimulate the students in actively participating in discussions raising questions and expressing personal opinions, in particular during the seminars. Each student will execute the experimental protocol (lab module) individually and will be stimulated to compare his(her) own experience with other students. At the end of the laboratory, each student has to write a report of the experience.

As concerns the teaching methods of this course unit, all students must attend Module 1, 2 [https://www.unibo.it/en/services-and-opportunities/health-and-assistance/health-and-safety/online-course-on-health-and-safety-in-study-and-internship-areas] online, while Module 3 on health and safety is to be attended in class. Information about Module 3 attendance schedule is available on the website of your degree programme.

Assessment methods

The exam will assess the knowledge level of each student of the mechanisms of genome integrity, DNA damage and repair, which are at the basis of human syndromes and chronic diseases such as cancer and neurodegeneration. The exam will also assess if the student is familiar with experimental protocols used during the lab practice.

The exam consists of: a) a written report at the end of the laboratory, and b) an oral exam. The oral exam will consist of questions related to the program of the course and a discussion of the written report of the laboratory.
Minimum score to pass the exam is 18 out of 30.

Teaching tools

Power point presentations, PC and internet. The laboratory is fully equipped with instruments and reagents needed for the planned experimental activity.

Office hours

See the website of Giovanni Capranico

See the website of Giulia Miglietta