STRUCTURAL
ORGANIZATION
OF
THE
MALARIA
MOSQUITO
HETEROCHROMATIN: FROM CHROMOSOMAL MORPHOLOGY TO GENOME
SEQUENCE
Maria V. Sharakhova1, Phillip George1, Irina V. Brusentsova2, Scotland Leman3,
Christopher D. Smith4, Igor V. Sharakhov1*
1
Department of Entomology, Virginia Tech, Blacksburg, VA, USA; 2 Department of
Molecular and Cellular Biology, Institute of Chemical Biology and Fundamental Medicine
SB RAS, Novosibirsk, Russia; 3Department of Statistics, Virginia Tech, Blacksburg, VA,
USA; 4Department of Biology, San Francisco State University, San Francisco, CA, USA;
e-mail:igor@vt.edu; *corresponding author
Key words: heterochromatin, malaria mosquito, imunostaning, Bayesian statistics
Motivations and aim: Heterochromatin plays an important role in chromosome function and
gene regulation. The goal of this study was to map and characterize the heterochromatic
portion of the malaria mosquito Anopheles gambiae genome.
Methods and Algorithms: Imunostaining of An. gambiae polytene chromosomes was
performed using antibodies against Heterochromatin Protein 1 (HP1) and lamin Dm0
Drosophila melanogaster. Physical mapping was done by Cy3, Cy5 labeled PCR amplified
genes. The bioinformatics analysis was conducted using C program ATCONTENT, Biomart
and SMARTest. We developed novel Bayesian statistical models to analyze genome
landscapes in different types of chromatin. A gene ontology (GO) annotation file was
generated using Interproscan-assigned GO terms and custom Perl scripts. Segmental
duplications were detected using BLAST-based whole-genome assembly comparison limited
to putative segmental duplications represented by pairwise alignments with ≥2.5-kb
and >90% sequence identity.
Results: Based on chromosomal location, two types of heterochromatin have been identified
in An. gambiae chromosomes: pericentromeric and intercalary. Intercalary heterochromatin
was represented by two morphological forms: compact, block-like type and diffuse, meshlike type. Pericentromeric heterochromatin in An. gambiae has a mesh-like structure.
Immunostaining of the An. gambaie chromosomes with antibodies against HP1 and lamin
Dm0 identified the major invariable sites of protein localization in regions of diffuse
heterochromatin and euchromatic region 9C but not in the compact intercalary
heterochromatin. To determine the euchromatin-heterochromatin boundaries in the An.
gambaie genome, we physically mapped genes to the polytene chromosomes: 16.6 Mb of the
mapped portion in the An. gambiae genome has been identified as heterochromatin. Unlike
Drosophila, Anopheles possesses three large regions (0.7, 0.8, and 2.9 megabase pairs long)
of intercalary heterochromatin. The study determined 230 in the mapped portion of the An.
gambiae heterochromatin. GO analysis revealed that heterochromatin is enriched by genes
with DNA-binding and regulatory activities. Bayesian statistical models showed that
heterochromatin and euchromatin differ in gene density and that all chromatin types are
distinguishable from each other by the coverage of transposable elements. Conversely,
segmental duplication, matrix-associated regions, and simple repeats are distributed
regardless of the chromatin type. Finally, we have demonstrated that the 42 Mb assembly of
unmapped scaffolds (“unknown” chromosome) have molecular characteristics of
heterochromatin.
Conclusion: The study revealed the presence of large blocks of intercalary heterochromatin in
An. gambiae genome. We conclude that each chromatin type has a unique combination of
various classes of genes and repetitive DNA.