Interaction between SWP9 and Polar Tube Proteins of the Microsporidian Nosema bombycis and Function of SWP9 as a Scaffolding Protein Contribute to Polar Tube Tethering to the Spore Wall

Abstract

All microsporidia possess a unique, highly specialized invasion mechanism that involves the polar tube and spore wall. The interaction between spore wall proteins (SWPs) and polar tube proteins (PTPs) in the formation, arrangement, orderly orientation, and function of the polar tube and spore wall remains to be determined. This study was undertaken to examine the protein interactions of Nosema bombycis SWP7 (NbSWP7), NbSWP9, and PTPs. Coimmunoprecipitation, liquid chromatographytandem mass spectrometry (LC-MS/MS), and yeast two-hybrid data demonstrated that NbSWP9, but not NbSWP7, interacts with NbPTP1 and NbPTP2. Furthermore, immunoelectron microscopy (IEM) showed that NbSWP9 was localized mainly in the developing polar tube of sporoblasts, while NbSWP7 was found randomly in the cytoplasm. However, both NbSWP9 and NbSWP7 were located in the polar tube and spore wall of N. bombycis mature spores. The reason why NbSWP7 was localized to the polar tube may be due to the interaction between NbSWP9 and NbSWP7. Interestingly, the majority of NbSWP9, but not NbSWP7, accumulated in the beginning part of the extruded polar tube and the ruptured spore wall called the anchoring disk (AD) when the mature spores germinated under weak-alkaline environmental stimulation. Additionally, anti-NbSWP9 antibody reduced spore germination in a dose-dependent manner. In conclusion, our study further confirmed that NbSWP9 is a scaffolding protein that not only anchors and holds the polar tube but also tethers the polar tube to the spore wall.

Comparative Analysis of the Proteins with Tandem Repeats from 8 Microsporidia and Characterization of a Novel Endospore Wall Protein Colocalizing with Polar Tube from Nosema bombycis

“…In this paper, we found that EOB14572 can also be detected in the beginning and end terminal part of the extruded polar tube in germinated spore, which is the same as NbSWP9 reported recently (Yang et al. ). Thus, we speculate EOB14572 might be a new component of polar tube or interact with polar tube proteins of N. bombycis .…”

“…IFA and IEM results demonstrated that the hypothetical protein EOB14572 (NBO_24g0018) was an endospore wall protein of N. bombycis, suggesting that this protein might contribute to the spore wall formation. More interestingly, three spore wall proteins NbSWP5, NbSWP7 and NbSWP9 were previously described that they could colocalize with the extruded polar tube of N. bombycis (Li et al 2012;Yang et al 2015Yang et al , 2017. In this paper, we found that EOB14572 can also be detected in the beginning and end terminal part of the extruded polar tube in Figure 3 Immunoelectron microscopy of EOB14572 in mature spores of Nosema bombycis.…”

“…; Yang et al. , ). In this paper, we found that EOB14572 can also be detected in the beginning and end terminal part of the extruded polar tube in germinated spore, which is the same as NbSWP9 reported recently (Yang et al.…”

A Novel Spore Wall Protein from Antonospora locustae (Microsporidia: Nosematidae) Contributes to Sporulation

“…, ; Yang et al. , , ). All of these spore wall proteins can be localized to the exospore, endospore, or plasmalemma, and nearly all of these have functional binding sites similar to the heparin‐binding motif (HBM), or modifications such as phosphorylation and glycosylation.…”

“…The microsporidian spore wall proteins identified so far, have very low or no sequence similarity with any other eukaryotic proteins: SWP1, SWP2, and EnP1 from Encephalitozoon intestinalis (Hayman et al 2001;Southern et al 2007); EnP1 from Encephalitozoon romaleae (Pombert et al 2012); EcSWP1, EcCDA, EnP1, EnP2, and SWP3 from Encephalitozoon cuniculi (Brosson et al 2005;Peuvel-Fanget et al 2006;Taupin et al 2006;Xu et al 2006); Ehswp1a and Ehswp1b from Encephalitozoon hellem (Polonais et al 2010); as well as NbSWP7, NbSWP9, NbSW16, NbHSWP11, SWP5, NbSWP12, SWP25, SWP26, SWP30, and SWP32 from Nosema bombycis (Chen et al 2013;Li et al 2009Li et al , 2012Wang et al 2015;Wu et al 2008Wu et al , 2009Yang et al 2014Yang et al , 2015Yang et al , 2017. All of these spore wall proteins can be localized to the exospore, endospore, or plasmalemma, and nearly all of these have functional binding sites similar to the heparin-binding motif (HBM), or modifications such as phosphorylation and glycosylation.…”