Cytotoxic effects of pyrrolizidine alkaloids

Pereira, Tamara Nishanthi. (2004). Cytotoxic effects of pyrrolizidine alkaloids PhD Thesis, School of Molecular and Microbial Sciences, The University of Queensland.

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Author Pereira, Tamara Nishanthi.
Thesis Title Cytotoxic effects of pyrrolizidine alkaloids
School, Centre or Institute School of Molecular and Microbial Sciences
Institution The University of Queensland
Publication date 2004
Thesis type PhD Thesis
Supervisor Dr. Arun S. Prakash
Dr. Simon Worrall
Total pages 203
Collection year 2004
Language eng
Subjects L
270199 Biochemistry and Cell Biology not elsewhere classified
780105 Biological sciences
Formatted abstract

Pyrrolizidine alkaloids (PAs) are a group of naturally occurring toxins found in flowering plants of widespread geographical distribution. Ingestion of PAs results in liver toxicity and such poisonings have been reported in humans and animals. The liver is the main target organ as this is the site of metabolic activation of the parent alkaloids to toxic constituents. Persistent, enlarged hepatocytes are a characteristic feature of PA poisoning, however the exact mechanism of hepatocyte enlargement is not well understood. This study was aimed at evaluating the hypothesis that such enlarged cells are the result of PA-induced DNA damage resulting in a mitotic block. 


In vitro studies showed that dehydromonocrotaline (DHM), a toxic metabolite of a commonly occurring PA monocrotaline, can modify DNA in several different ways. DHM was shown to modify DNA at the N-7 of guanine (G) in the major groove with a preference for guanines in 5'-GG and 5'-GA sequences (where A represents adenine). DHM is a bi-functional alkylating agent which can form inter-strand DNA cross-links. In addition to the two electrophilic sites, DHM also has a nucleophilic site and can polymerise with itself. Such DHM polymers can form inter-fragment DNA cross-links involving several DNA fragments. These cross-links are believed to involve the N-2 of G's in the minor groove of DNA. It is conceivable that all these forms can occur in vivo and be of potential biological significance. Four in vivo candidate systems were evaluated for their ability to produce PA-induced enlarged cells in a reliable manner. The yeast S. cerevisae and human myeloid leukaemia (HL60) systems were found to be unsuitable as these cells did not express the enzymes needed to convert parent alkaloids to toxic constituents. The chicken embryo system was also unsuitable because liver damage was too severe in this model. The weanling rat model was shown to be the most suitable one for studying PA-induced hepatic megalocytosis. While rats are small animals with high metabolic rates and short life-spans, they appear to develop PA-induced liver disease in a similar manner to humans. They are susceptible to PAs by the oral route (mimicking human ingestion), have the relevant enzymes to convert PAs to toxic metabolites and develop enlarged hepatocytes characteristic of PA toxicity. PA-induced DNA damage was investigated further in the hepatic rat gene for p53 (TP53). A mutation was seen in TP53 from liver tissue of rats with the most severe hepatocyte cell enlargement. The mutation seen was at codon 152/153 in exon 5 of TP53 and was an T to A transversion of the last base of codon 152 and the first base of codon 153 (GGT ACC to GGA TCC). 


Based on these results a schema is proposed for PA-induced hepatocyte enlargement. This study has shown that the PA monocrotaline results in a mutation from ACC (threonine) to TCC (serine) at codon 153 of the rat TP53. This corresponds to codon 155 of the human TP53 and this amino acid is normally phosphorylated by the protein kinase CK2. The mutation may alter the phosphorylation of the p53 protein by CK2 such that the p53/mdm2 interaction is enhanced, facilitating the degradation of p53 by the ubiquitin-26S proteosome system. The resultant decrease in p53 protein means it cannot fulfil its function as a transcription factor for downstream genes. One such gene is p21Waf 1/Cip 1/Sdi 1 and cells lacking functional p21 have been shown to be in a mitotic block.   

Keyword Pyrrolizidines
Poisonous plants

Document type: Thesis
Collection: UQ Theses (RHD) - UQ staff and students only
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Created: Fri, 24 Aug 2007, 18:36:17 EST