[Background] The tick-borne intracellular pathogen, Anaplasma phagocytophilum (Rickettsiales: Anaplasmataceae) causes human granulocytic anaplasmosis after infection of polymorphonuclear leucocytes. The human Sp110 gene is a member of the nuclear body (NB) components that functions as a nuclear hormone receptor transcriptional coactivator and plays an important role in immunoprotective mechanisms against pathogens in humans. In this research, we hypothesized that Sp110 may be involved in the infection of human promyelocytic HL-60 cells with A. phagocytophilum.
[Methods] The human Sp110 and A. phagocytophilum msp4 mRNA levels were evaluated by realtime
RT-PCR in infected human HL-60 cells sampled at 0, 12, 24, 48, 72 and 96 hours postinfection.
The effect of Sp110 expression on A. phagocytophilum infection was determined by RNA
interference (RNAi). The expression of Sp110 was silenced in HL-60 cells by RNAi using predesigned
siRNAs using the Nucleofector 96-well shuttle system (Amaxa Biosystems, Gaithersburg, MD, USA). The A. phagocytophilum infection levels were evaluated in HL-60 cells after RNAi by realtime PCR of msp4 and normalizing against human Alu sequences.
[Results] While Sp110 mRNA levels increased concurrently with A. phagocytophilum infections in HL-60 cells, the silencing of Sp110 expression by RNA interference resulted in decreased infection levels.
[Conclusion] These results demonstrated that Sp110 expression is required for A. phagocytophilum infection and multiplication in HL-60 cells, and suggest a previously undescribed mechanism by which A. phagocytophilum modulates Sp110 mRNA levels to facilitate establishment of infection of
human HL-60 cells.
This work was supported by the Oklahoma Agricultural Experiment Station
(project 1669), the Sitlington Endowed Chair for Food Animal Research to
K.M.K. and the Ministry of Science and Education (MEC), Spain (project
AGL2005-07401). Dr. Raúl Manzano-Roman was funded by Ministerio de
Educación y Ciencia, Spain. V. Naranjo was founded by Consejería de Educación,
JCCM, Spain.
Peer reviewed