R1 Region of P97 Mediates Adherence ofMycoplasma hyopneumoniaeto Swine Cilia

Abstract

Adherence of Mycoplasma hyopneumoniae to the swine respiratory tract is mediated by the membrane protein P97. This protein is located on the outer membrane surface, and its role in adherence has been firmly established. The general region of P97 that mediates adherence to swine cilia is thought to be the R1 region near the carboxy terminus of the protein, but it was not clear if this region could mediate adherence to swine cilia independently of other P97 sequences. To examine this in more detail, a series of R1 repeat sequences containing different numbers of repeating units cloned in frame with lacZ was used to produce R1-␤-galactosidase fusion proteins. These proteins were then tested for adherence to swine cilia and for reactivity to the adherence-blocking monoclonal antibody F2G5 and convalescent-phase swine sera. In this way it was possible to accurately define the cilium binding epitope of P97 and the minimal epitope recognized by antibody. Our results indicate that eight R1 repeating units are required for cilium binding and that three repeating units are needed for antibody recognition. These results could lead to more effective therapeutic measures against this important swine pathogen.Attachment to host tissue is essential for colonization by most mucosal pathogens, including mycoplasmas (1, 2). The best-defined mycoplasmal adhesins, those of Mycoplasma pneumoniae and Mycoplasma genitalium, are found in a specialized attachment organelle (8). It is clear from numerous studies that their adhesins work in concert with other membrane proteins and accessory factors in a coordinated fashion to form the attachment organelle structure (8). Thus, the process of adherence in these species is multifactorial, requiring both membrane proteins and cytoskeletal elements (8). Most mycoplasmas, however, lack specialized attachment organelles like M. pneumoniae, and consequently, their adherence mechanisms differ. How most mycoplasma species adhere to their respective host tissues is largely unknown, and the genetic mechanisms of adherence are poorly defined.As the etiological agent of porcine mycoplasma pneumonia and a major component of the porcine respiratory disease complex, Mycoplasma hyopneumoniae continues to present significant problems for the swine industry. As is the case for many other mycoplasmal species, it is host specific (for swine), it has no readily apparent external structures or organelles that could be used for attachment, there are no genetic systems for analyzing the adherence phenotype, and studies of adherence have been largely qualitative rather than quantitative in nature. For instance, M. hyopneumoniae closely adheres to swine respiratory epithelium both in vivo and in vitro (3, 9). It has also been demonstrated that M. hyopneumoniae can attach to pig and human lung fibroblasts, to pig kidney cells in culture (15), and to turkey red blood cells (12).Our interests in adherence mechanisms as the prelude to colonization and disease led us to take advantage of the available adherence-blocking ...

P159 is a proteolytically processed, surface adhesin ofMycoplasma hyopneumoniae: defined domains of P159 bind heparin and promote adherence to eukaryote cells

“…These advances are significant given that overall poor growth on agar surfaces coupled with a lack of development of genetic tools to selectively mutate target genes has for many years hampered efforts to identify molecules that play fundamental roles in pathogenesis (Minion, 2002). The cilium adhesin, P97 is the only cell surface adhesin that has been extensively characterized in M. hyopneumoniae (Hsu and Minion, 1998;Minion et al ., 2000). P97 is extensively cleaved post-translationally during growth in vitro and processing is strain-specific.…”

Etiopatogenia e imunoprofilaxia da pneumonia enzoótica suína

“…Lipoproteínas, que são os principais componentes de membrana de micoplasmas, compreendem aproximadamente 2/3 da massa total da membrana (RAZIN et al, 1998) e são elas que normalmente sofrem uma variação antigênica frente à resposta imune do hospedeiro. Por outro lado, a função dos genes homólogos à adesina P97 é desconhecida e provavelmente não está relacionada com a adesão, uma vez que apenas um deles possui o domínio repetitivo R1 responsável pela ligação ao receptor ciliar (MINION et al, 2000;MINION et al, 2004).…”

“…Cepas de M. hyopneumoniae com variação no número de repetições da região R1 (8 a 15) foram identificadas (WILTON et al, 1998), sugerindo que esta variação poderia influenciar a capacidade de ligação ao cílio suíno. Porém, MINION et al (2000), utilizando fusões de β-galactosidase com diferentes números de repetições R1, concluíram que no mínimo oito repetições são requeridas para a ligação ao cílio suíno e três para o reconhecimento dos MAbs Ciência Rural, v.36, n.3, mai-jun, 2006.…”

Association of number of tandem repeats in two important adhesins in Mycoplasma hyopneumoniae

“…Repeat region 1 (RR1), a locus containing tandem repeats (TR) in the gene mhp183 (P97) has been suggested as to be immunogenic and essential for ciliary binding (Wilton et al, 1998;Minion et al, 2000). The TR in RR1 can vary between strains and this variability can influence the ciliary binding (Wilton et al, 1998).…”

“…The TR in RR1 can vary between strains and this variability can influence the ciliary binding (Wilton et al, 1998). At least eight repeats in RR1 have been suggested as needed for cilium binding, and three repeats are required for antibody recognition (Minion et al, 2000). In the gene mhp 684 (P146), the locus of the polyserine RR3 is of unknown function, although it is located in a P85 fragment that is associated with binding to porcine respiratory cilia and plasminogen (Bogema et al, 2012).…”