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C Frontali

Publications and source records attributed to C Frontali.

At least 37 records · Page 2Linked to original sources

Long insertions within telomeres contribute to chromosome size polymorphism in Plasmodium berghei.

During prolonged in vivo mitotic multiplication of a Plasmodium berghei ANKA clone (8417HP), parasites that contained an enlarged version of chromosome 4 were observed. Restriction mapping and hybridization results demonstrated that the extra DNA present in the enlarged chromosome consists of 2.3-kb tandem repeats, known to be normally located in subtelomeric position at several chromosomal ends but absent in the original chromosome. The inserted 2.3-kb units appeared to interrupt one of the original telomeres and to create an internal (approximately 1-kb-long) telomeric sequence.

Animals↗

Organization of subtelomeric repeats in Plasmodium berghei.

Several (but not all) Plasmodium berghei chromosomes bear in the subtelomeric position a cluster of 2.3-kilobase (kb) tandem repeats. The 2.3-kb unit contains 160 base pairs of telomeric sequence. The resulting subtelomeric structure is one in which stretches of telomeric sequences are periodically spaced by a 2.1-kb reiterated sequence. This periodic organization of internal telomeric sequences might be related to chromosome-size polymorphisms involving the loss or addition of subtelomeric 2.3-kb units.

Animals↗

Telomeric motifs are present in a highly repetitive element in the Plasmodium berghei genome.

Using as probes the subfragments of the telomeric sequence previously cloned by us from Plasmodium berghei DNA, we identified and cloned a 2.3 kb repeat, largely overlapping the original telomeric insert. Restriction mapping indicated that cloned inserts (2.3 kb in length) represented circularly permutated versions of a rather well conserved repeated element, at least in part organized in tandem. The 2.3 kb repeat family with a copy number of about 300 occupies about 4% of the whole genome. The copies are unevenly distributed among the chromosome-sized molecules revealed by pulsed field gradient electrophoresis. Complete sequence determination of the 2.3 kb element revealed that telomere-related motifs are present with a characteristic pattern in a set of tandem repeats, 27 bp long. The perfect conservation of these motifs as well as the pattern of chromosomal distribution suggest that we are dealing with a specialised structure subject to selective mechanisms of amplification and maintenance.

Animals↗

Homologous telomeric sequences are present in different species of the genus Plasmodium.

The telomeric sequence cloned from Plasmodium berghei (see M. Ponzi et al. (1985) EMBO J. 4, 2991-2995) was tested for species specificity. A telomeric and a subtelomeric fragment of the cloned insert served as separate, labelled probes on pulsed field gradient electrophoretical patterns and on genomic digests from the rodent malarias Plasmodium yoelii, Plasmodium chabaudi and from the human malaria Plasmodium falciparum. Results indicate that the subtelomeric fragment, abundantly represented in two chromosomes of P. berghei, is not present in the other DNA tested, while the telomeric fragment is present in every chromosome-sized molecule in all the species tested. The telomeric location in the other genomes of the sequences homologous to the P. berghei telomeric probe is confirmed by experiments with Bal 31 exonuclease. In all cases, the TaqI site appears to delimit the common telomeric portion.

Animals↗

Identification of a telomeric DNA sequence in Plasmodium berghei.

A fragment of Plasmodium berghei DNA was cloned using a technique designed to select for telomeric sequences. The cloned fragment recognizes Bal31-sensitive bands in P. berghei genomic digests. It contains at its distal end at least 70 tandem repeats of the heptanucleotide sequence CCCTGAAA. The presence of natural single strand discontinuities in the telomeric regions of P. berghei DNA is demonstrated by the selective incorporation of deoxyribonucleoside triphosphates in the absence of DNase. The number of copies of the cloned sequence present in each genome agrees with an estimate of 6-12 chromosomes per nucleus.

Animals↗

Re-examination of earlier work on repetitive DNA and mosquito infectivity in rodent malaria.

Previous results, relating mosquito infectivity to percentage of repetitive DNA in the genome of Plasmodia, are re-examined in the light of the finding that a parasite line used in the previous studies and classified as Plasmodium berghei NK65, was a mixed infection, where the major component appeared to be Plasmodium yoelii. This conclusion was reached through cloning and isoenzyme typing of different clones. Isoenzyme typing alone is not sufficiently sensitive to reveal contamination amounting to less than 20% in a mixture. Attention is drawn to the risk inherent in work with uncloned lines, where the proportions of species or sub-species present may vary according to line history and gametocyte viability.

Animals↗

Further studies and electron microscopic characterization of Plasmodium berghei DNA.

The average length and the interspersion pattern of repetitive DNA sequences in the Plasmodium berghei genome have been studied by electron microscopy. Within the limitations posed by the relatively high genome complexity, analysis of partially renatured total DNA indicates that repetitive sequences do not occupy preferential positions along the genome, but are widely dispersed (one in approx. 8000 base pairs of unique DNA). Structures appearing as loops flanked by inverted repeats are present. Analysis of the repetitive fraction purified by hydroxyapatite chromatography shows that the average length of rapidly reassociating repetitive structures is around 800 base pairs with 90% of the length distribution between 400 and 1400 base pairs. Suitable extraction methods, preserving circularity of extrachromosomal DNA components, allow the detection of molecules which can be identified as mitochondrial DNA, 10.5 +/- 0.4 microns long.

Animals↗

Mosquito infectivity is directly related to the proportion of repetitive DNA in Plasmodium berghei.

A strain of Plasmodium berghei (NK 65) was followed during syringe transmission in mice for over 120 passages after the last complete cycle, while the following parameters were monitored: (a) capacity to infect mosquitoes, inducing oocyst formation; (b) presence in the peripheral blood of morphologically identifiable gametocytes; (c) presence of a repetitive component in the DNA extracted from intraerythrocytic population. The suggestion of a possible role of this component in gametogenesis came from an earlier work (Dore, E., Birago, C., Frontali, C. and Battaglia, P.A. (1980) Mol. Biochem. Parasitol. 1, 199-208). Present results confirm the correlation between proportion of repetitive DNA and infectivity towards mosquitoes with a correlation coefficient r = 0.92-0.07+0.04. A parallel decrease of the two quantities is observed in the course of syringe transmission. A limited number of cloned lines, derived from strain NK 65 at different times during syringe transmission, shared the infectivity properties of the parent strain at the moment of cloning, thus confirming that in the infective stage single asexual parasites from the schizogonic cycle are able to originate the whole cycle. The above arguments and results suggest that differentiation into active gametocytes involves amplification of a portion of the genome.

Aedes↗

[Calibration of gel-chromatographic columns for examination of dextrans for clinical use].

A procedure is described which allows to obtain integral distribution in molecular weights for oligodextran preparations of clinical use from the elution profiles obtained by gel-filtration. The calibration procedure requires the use of commercially available dextran fractions with known distribution in molecular weight (the M.W. distribution of the standards may be checked through absolute determination of Mn and Mw). For given column dimensions and elution conditions, the outlined procedure yields an accurate plot of log Mw vs k (partition coefficient).

Chromatography, Gel↗