Ca2+-mediated exocytosis of subtilisin-like protease 1: a key step in egress of Plasmodium falciparum merozoites
Version of Record online: 28 DEC 2012
© 2012 John Wiley & Sons Ltd
Volume 15, Issue 6, pages 910–921, June 2013
How to Cite
Agarwal, S., Singh, M. K., Garg, S., Chitnis, C. E. and Singh, S. (2013), Ca2+-mediated exocytosis of subtilisin-like protease 1: a key step in egress of Plasmodium falciparum merozoites. Cellular Microbiology, 15: 910–921. doi: 10.1111/cmi.12086
- Issue online: 14 MAY 2013
- Version of Record online: 28 DEC 2012
- Accepted manuscript online: 6 DEC 2012 06:36AM EST
- Manuscript Accepted: 23 NOV 2012
- Manuscript Revised: 30 OCT 2012
- Manuscript Received: 14 AUG 2012
- Innovative Young Biotechnology Award (IYBA)
- European Commission
Videos S1–S4. Intracellular Ca2+ levels during egress of P. falciparum merozoites from host erythocytes. P. falciparum late-stage schizonts were labelled with Fluo-4AM and added to erythrocytes. Both DIC and fluorescence images were acquired simultaneously on a Zeiss LSM510 confocal microscope with a 1.4 NA lens, using 488 nm excitation at low power to minimize bleaching and cell damage. Videos S1A, S2A, S3A, S4A: merge of the DIC and fluorescence images are shown. Videos S1B, S2B, S3B, S4B: DIC images are shown. Videos S1C, S2C, S3C, S4C: fluorescence images are shown.
Fig. S1. A–C. Analysis of intracellular Ca2+ levels during egress of P. falciparum merozoites. Late-stage purified P. falciparum schizonts were labelled with Fluo-4AM and added to uninfected erythrocytes. Intracellular Ca2+ levels were monitored in P. falciparum schizonts during egress by time-lapse video microscopy. Selected pairs of merge of both DIC and fluorescence images are shown. Mean fluorescence intensities (MFI) after background subtraction is reported for schizonts from three independent experiments corresponding to Videos S2, S3 and S4 from which merozoites were released following its rupture. An increase in intracellular Ca2+ just before egress was observed in each schizonts.
Fig. S2. Expression and purification of the recombinant rPfSERA5 and rPfSUB1 proteins. The amino acid region (E578 to N828, within catalytic domain) of PfSERA5 and amino acid region (E27 to H688, full-length excluding signal sequence) of PfSUB1 was expressed and purified to homogeneity.
A. The purified protein of PfSERA5 was analysed on a SDS-PAGE and detected in immunoblot using anti-His tag antibody.
B. The purified protein of PfSUB1 was analysed on a SDS-PAGE and detected in immune-blots using an anti-His tag antibody.
Fig. S3. Western blot analysis to confirm the specificity of anti-PfSUB1 mouse sera. Western blot of lysates prepared from ring-infected erythrocytes (∼ 14–16 hpi) and schizont-infected erythrocytes (∼ 48 hpi) as well as of uninfected erythrocytes, with anti-PfSUB1 mouse sera. Mature 47 kDa along with 54 kDa fragment of PfSUB1 detected only in schizont-infected erythrocytes lysate.
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