Intron-containing beta-tubulin transcripts in Cryptosporidium parvum cultured in vitro.
The genome of Cryptosporidium parvum contains a relatively small number of introns, which includes the beta-tubulin gene with only a single intron. Recently, it was observed that the intron was not removed from some of the beta-tubulin transcripts in the late life cycle stages cultured in vitro. Although normally spliced beta-tubulin mRNA was detected in all parasite intracellular stages by RT-PCR (e.g. HCT-8 or Caco-2 cells infected with C. parvum for 12-72 h), at 48-72 h post-infection unprocessed beta-tubulin transcripts containing intact introns started to appear in parasite mRNA within infected host cells. The intron-containing transcripts could be detected by fluorescence in situ hybridization (FISH) using an intron-specific probe. The intron-containing beta-tubulin transcripts appeared unique to the in vitro-cultured C. parvum, since they were not detected in parasite-infected calves at 72 h. As yet, it is unclear whether the late life cycle stages of C. parvum are partially deficient in intron-splicing or the intron-splicing processes have merely slowed, both of which would allow the detection of intron-containing transcripts. Another possible explanation is that the decay in transcript processing might simply be due to the onset of parasite death. Nonetheless, the appearance of intron-containing transcripts coincides with the arrest of C. parvum development in vitro. This unusual observation prompts speculation that the abnormal intron-splicing of beta-tubulin transcripts may be one of the factors preventing complete development of this parasite in vitro. Furthermore, the presence of both processed and unprocessed introns in beta-tubulin transcripts in vitro may provide a venue for studying overall mechanisms for intron-splicing in this parasite.
Cai
Xiaomin
X
Department of Veterinary Pathobiology, College of Veterinary Pathobiology, Texas A&M University, 4467 TAMU, College Station, TX 77843-4467, USA.
Lancto
Cheryl A
CA
Department of Veterinary Pathobiology, University of Minnesota, St Paul, MN 55108, USA.
Abrahamsen
Mitchell S
MS
Department of Veterinary Pathobiology, University of Minnesota, St Paul, MN 55108, USA.
Zhu
Guan
G
Faculty of Genetics Program, Texas A&M University, 4467 TAMU, College Station, TX 77843-4467, USA.|Department of Veterinary Pathobiology, College of Veterinary Pathobiology, Texas A&M University, 4467 TAMU, College Station, TX 77843-4467, USA.
D000818:Animals; D018938:Caco-2 Cells / Q000469:parasitology; D002417:Cattle; D002418:Cattle Diseases / Q000469:parasitology; D002460:Cell Line; D003457:Cryptosporidiosis / Q000469:parasitology / Q000662:veterinary; D016785:Cryptosporidium parvum / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007438:Introns; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D014404:Tubulin / Q000235:genetics* / Q000378:metabolism*
D012333:RNA, Messenger; D014404:Tubulin