Knockout of the gene encoding the kinetoplast-associated protein 3 (KAP3) in Trypanosoma cruzi: effect on kinetoplast organization, cell proliferation and differentiation.
Kinetoplast DNA (kDNA) of trypanosomatid protozoa consists of an unusual arrangement of two types of circular molecules catenated into a single network: (1) a few maxicircles that encode various mitochondrial enzyme subunits and rRNA in a cryptic pattern and (2) thousands of minicircles that encode guide RNAs (gRNAs). kDNA is associated with proteins, known as kinetoplast-associated proteins (KAPs), which condense the kDNA network. However, little is known about the KAPs of Trypanosoma cruzi, a parasite that displays different kDNA condensation patterns during its complex morphogenetic development. We cloned the T. cruzi gene encoding TcKAP3 (kinetoplast-associated protein 3). TcKAP3 is a single-copy gene that is transcribed into a 1.8-kb mRNA molecule and expressed in all stages of the parasite. Mouse antiserum raised against recombinant TcKPA3 recognized a 21.8kDa protein, which was found, by indirect immunofluorescence and immunoelectron microscopy, to be associated with the T. cruzi kinetoplast. Several features of TcKAP3, such as its small size, basic nature and similarity with KAP3 from the insect trypanosomatid Crithidia fasciculata, are consistent with a role in DNA charge neutralization and condensation. This suggests that this protein is involved in organizing the kDNA network. Gene deletion was used to investigate TcKAP3 function. Here we investigated the T. cruzi KAP3 null mutant, analyzing its fitness during proliferation, differentiation and infectivity.
de Souza
Flavia Sá Pereira
FS
Instituto Carlos Chagas, Fundação Oswaldo Cruz, Paraná, Brazil.
Rampazzo
Rita de Cássia Pontello
Rde C
Manhaes
Lauro
L
Soares
Maurilio José
MJ
Cavalcanti
Danielle Pereira
DP
Krieger
Marco Aurélio
MA
Goldenberg
Samuel
S
Fragoso
Stenio Perdigão
SP
D003001:Cloning, Molecular; D018105:DNA, Kinetoplast / Q000378:metabolism*; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D055786:Gene Knockout Techniques; D046911:Macromolecular Substances / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D014349:Trypanosoma cruzi / Q000235:genetics / Q000254:growth & development / Q000502:physiology*
D018105:DNA, Kinetoplast; D016054:DNA, Protozoan; D046911:Macromolecular Substances; D015800:Protozoan Proteins; C064816:kinetoplast-associated protein, protozoan