Unraveling the role of cAMP signaling in : insights into PKA-mediated regulation of encystation and subcellular interactions.

Unlabelled: cAMP plays an important role as a second messenger in the stage transition of various protozoan parasites. This signaling pathway relies on multiple effectors, such as protein kinase A (PKA), exchange protein activated by cAMP, and cAMP-response element binding protein transcription factors, to initiate signal transduction in humans. The genome only contains two adenylate cyclases (ACs), a single phosphodiesterase (PDE) and a single known PKA effector, and the specific functions of these components are not fully understood.

Encystation stimuli sensing is mediated by adenylate cyclase AC2-dependent cAMP signaling in Giardia.

Protozoan parasites use cAMP signaling to precisely regulate the place and time of developmental differentiation, yet it is unclear how this signaling is initiated. Encystation of the intestinal parasite Giardia lamblia can be activated by multiple stimuli, which we hypothesize result in a common physiological change. We demonstrate that bile alters plasma membrane fluidity by reducing cholesterol-rich lipid microdomains, while alkaline pH enhances bile function.

Encystation stimuli sensing mediated by adenylate cyclase AC2-dependent cAMP signaling in .

Protozoan parasites use cAMP signaling to precisely regulate the place and time of developmental differentiation, yet it is unclear how this signaling is initiated. Encystation of the intestinal parasite can be activated by multiple stimuli, which we hypothesize result in a common physiological change. We demonstrate that bile alters plasma membrane fluidity by reducing cholesterol-rich lipid microdomains, while alkaline pH enhances bile function.

Encystation stimuli sensing mediated by adenylate cyclase AC2-dependent cAMP signaling in .

Protozoan parasites use cAMP signaling to precisely regulate the place and time of developmental differentiation, yet it is unclear how this signaling is initiated. Encystation of the intestinal parasite can be activated by multiple stimuli, which we hypothesize result in a common physiological change. We demonstrate that bile alters plasma membrane fluidity by reducing cholesterol-rich lipid microdomains, while alkaline pH enhances bile function.

A cell-cycle-dependent GARP-like transcriptional repressor regulates the initiation of differentiation in .

Transcriptional regulation of differentiation is critical for parasitic pathogens to adapt to environmental changes and regulate transmission. In response to encystation stimuli, Giardia lamblia shifts the distribution of the cell cycle toward G2 and induces the expression of cyst wall proteins (CWPs) within 2 to 4 h, indicating that key regulatory steps occur within the first 4 h of encystation. However, the role of transcription factors (TFs) in encystation has primarily been investigated at later time points.

The Giardia ventrolateral flange is a lamellar membrane protrusion that supports attachment.

Attachment to the intestinal epithelium is critical to the lifestyle of the ubiquitous parasite Giardia lamblia. The ventrolateral flange is a sheet-like membrane protrusion at the interface between parasites and attached surfaces. This structure has been implicated in attachment, but its role has been poorly defined.

Staging Encystation Progression in Using Encystation-Specific Vesicle Morphology and Associating Molecular Markers.

Differentiation into environmentally resistant cysts is required for transmission of the ubiquitous intestinal parasite . Encystation in requires the production, processing and transport of Cyst Wall Proteins (CWPs) in developmentally induced, Golgi-like, Encystation Specific Vesicles (ESVs). Progress through this trafficking pathway can be followed by tracking CWP localization over time.

Nek8445, a protein kinase required for microtubule regulation and cytokinesis in .

has 198 Nek kinases whereas humans have only 11. has a complex microtubule cytoskeleton that includes eight flagella and several unique microtubule arrays that are utilized for parasite attachment and facilitation of rapid mitosis and cytokinesis. The need to regulate these structures may explain the parallel expansion of the number of Nek family kinases.

Hybrid Structured Illumination Expansion Microscopy Reveals Microbial Cytoskeleton Organization.

Recently developed tissue-hydrogel methods for specimen expansion now enable researchers to perform super-resolution microscopy with ∼65 nm lateral resolution using ordinary microscopes, standard fluorescent probes, and inexpensive reagents. Here we use the combination of specimen expansion and the optical super-resolution microscopy technique structured illumination microscopy (SIM) to extend the spatial resolution to ∼30 nm. We apply this hybrid method, which we call ExSIM, to study the cytoskeleton of the important human pathogen Giardia lamblia including the adhesive disc and flagellar axonemes.

14-3-3 Regulates Actin Filament Formation in the Deep-Branching Eukaryote .

The phosphoserine/phosphothreonine-binding protein 14-3-3 is known to regulate actin; this function has been previously attributed to sequestration of phosphorylated cofilin. 14-3-3 was identified as an actin-associated protein in the deep-branching eukaryote ; however, lacks cofilin and all other canonical actin-binding proteins (ABPs). Thus, the role of 14-3-3 (Gl14-3-3) in actin regulation was unknown.

Myosin-independent cytokinesis in utilizes flagella to coordinate force generation and direct membrane trafficking.

Devoid of all known canonical actin-binding proteins, the prevalent parasite uses an alternative mechanism for cytokinesis. Unique aspects of this mechanism can potentially be leveraged for therapeutic development. Here, live-cell imaging methods were developed for to establish division kinetics and the core division machinery.