The Decrease in FlaA Observed in a flaB Mutant of Borrelia burgdorferi Occurs Posttranscriptionally

Abstract

The Lyme disease bacterium Borrelia burgdorferi is a motile spirochete with a flat-wave morphology. The periplasmic flagella, which are situated between the outer membrane sheath and cell cylinder, are essential for both the cell's wavy shape and motility. Here we focus on the structure and regulation of its periplasmic flagella. Previous studies have suggested that the periplasmic flagella consist of a polymer of the major filament protein FlaB and a minor protein, FlaA. We used immunoprecipitation methodology to present further evidence that FlaA is indeed a flagellar protein. In addition, in contrast to FlaA of the spirochete Brachyspira hyodysenteriae, B. burgdorferi FlaA did not impact the overall helical shape of the periplasmic flagella. We have previously shown that B. burgdorferi lacks the sigma factor-dependent cascade control of motility gene transcription found in other bacteria. To begin to understand motility gene regulation in B. burgdorferi, we examined the effects of an insertion mutation in flaB on the amounts of proteins encoded by motility genes. Of several motility gene-encoded proteins examined, only the amount of FlaA was decreased in the flaB mutant; it was 13% compared to the wild-type amount. Real-time reverse transcriptase PCR analysis indicated that this inhibition was not the result of a decrease in flaA mRNA. In addition, protein stability analysis suggested that FlaA was turned over in the flaB mutant. Our results indicate that the lack of FlaB negatively influences the amount of FlaA found in the cell and that this effect is at the level of either translational control or protein turnover.Borrelia burgdorferi is a motile spirochete that is the causative agent of Lyme disease. The organelles for motility, the periplasmic flagella (PFs), reside between the outer membrane sheath and protoplasmic cell cylinder. Approximately 7 to 11 PFs are subterminally attached at each end of the cell, and these filaments overlap in the center of the cell (4, 27, 31). Motility is likely to be an important virulence factor for these spirochetes, as B. burgdorferi penetrates into tissues where other organisms fail to invade (11,34). Moreover, motility has been postulated to be essential for the life cycle of these spirochetes in both the tick and the mammalian hosts (45, 55).We are beginning to understand the dynamics of B. burgdorferi motility (see references 11, 42, and 45 for recent reviews). Recent results indicate that the PFs have both skeletal and motility functions (50, 61). Cells of mutants that are targeted in flaB, which encodes the major PF protein, are nonmotile and are rod shaped instead of being a flat wave as are wild-type cells (50, 61). During translational motility, backward moving waves are generated that are responsible for cell displacement (28). These waves result from the rotation of the relatively rigid left-handed helically shaped PFs closely juxtaposed to and interacting with the flexible protoplasmic cell cylinder: the PFs act as a gear generating waves along the cell cylin...

Overexpression of CsrA (BB0184) Alters the Morphology and Antigen Profiles ofBorrelia burgdorferi

“…At this time, it is not known how the level of FlaB is regulated by borrelial CsrA, which could be either in conjunction with other mechanisms or through independent regulation of its levels (23,57). It appears that the levels of CsrA are variable in different isolates, and it is interesting to speculate that the levels of the 41-kDa protein corresponding to FlaB as observed on the Coomassie blue-stained gel (Fig.…”

“…This differential regulation could be significant in microenvironments such as the skin of a mammalian host where increased expression of adhesins such as DbpA or BBK32 may facilitate greater colonization capability in conjunction with reduced motility. The reduced levels of CsrA, favoring increased expression of FlaB, may provide an added advantage to the spirochetes under conditions requiring increased motility with a concomitant decrease in adherence (56,57). This initial characterization of the role of CsrA in regu-lating gene expression will help facilitate a better understanding of the mechanism and role of this potential global regulator in controlling the levels of pathophysiological determinants of B. burgdorferi that contribute to its survival and virulence in diverse hosts.…”

Borrelia burgdorferi Uniquely Regulates Its Motility Genes and Has an Intricate Flagellar Hook-Basal Body Structure

“…Polyclonal anti-recombinant CheW3 was provided by C. Li (unpublished data). The amounts of immunoreactive protein in the gels were determined using FluorChem spot densitometry as previously described (49). To analyze the stability of the FlgE high-molecular-weight complex, lysates were incubated for 60 min 23°C with either 8 M urea or 8 M guanidine-HCl.…”

“…Quantitative reverse transcriptase PCR (qRT-PCR) analysis was carried out as previously described (49). Briefly, total RNA was isolated using an RNeasy kit (Qiagen) and treated with DNase (Turbo DNase I; Ambion) to remove contaminating DNA.…”

Profile of patients with Baggio-Yoshinari Syndrome admitted at "Instituto de Infectologia Emilio Ribas"

“…Mutant Borrelias with genetic changes at its different genes has been previously described by MALAWISTA et al 23 , SADZIENE et al 31,32,33 , FINGERLE et al 14 , MOTALEB et al 28 , CHARON et al 9 . The existence of mutant spirochete expressed at atypical cystic morphologies, possibly without periplasmic flagella and decreased expression of outer surface proteins (Osp), justify all the particularities found in BYS.…”