The ER Protein Translocation Channel Subunit Sbh1 Controls Virulence of Cryptococcus neoformans

Santiago-Tirado, Felipe H.; Hurtaux, Thomas; Geddes‐McAlister, Jennifer; Nguyen, Duy; Helms, Volkhard; Doering, Tamara L.; Römisch, Karin

doi:10.1128/mbio.03384-22

Cited by 2 publications

(4 citation statements)

References 71 publications

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“…4C). Since the epitope for anti-Sbh1 (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23) does not include the S3/T5 phosphorylation sites and wildtype Sbh1 is >90% phosphorylated at steady state, this confirms that the phosphorylation of Sbh1 at S3…”

Section: Sbh1 S3/t5 Phosphorylation Results In a Conformational Chang...mentioning

confidence: 76%

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The conserved membrane-proximal domain of Sbh1/ Sec61β guides signal peptides into the Sec61 channel

Yadhanapudi,

Bhadra,

Barbieri

et al. 2024

Preprint

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Protein secretion begins with protein translocation through the universally conserved Sec61 channel in the endoplasmic reticulum (ER) membrane. The function of its β-subunit, called Sbh1 in yeast, remains poorly understood. Sbh1/Sec61β has a cytosolic domain consisting of two parts: the N-terminal approximately 40 amino acids are intrinsically unstructured, followed by a conserved membrane proximal (CMP) domain. Sbh1 is anchored to the Sec61 channel by a single C-terminal transmembrane domain. Phosphorylation of the Sbh1/Sec61β intrinsically unstructured domain allows regulated import of specific proteins into the ER. We show here that S3/T5 phosphorylation of Sbh1 results in a conformational change in the Sbh1 intrinsically unstructured N-terminus. We also address the function of the Sbh1/Sec61β CMP domain. We identify a potential interaction site in the Sbh1 CMP domain with the Sec61 N-terminal amphipathic helix using molecular dynamics (MD) modelling. Peptide panning revealed that the Sbh1 CMP domain contains a binding site for Sbh1-dependent signal peptides. Using site-directed mutagenesis, we show that the CMP region controls import of Sbh1-dependent translocation substrates into the ER. We conclude that the Sbh1 CMP domain functions as a gatekeeper that recognizes and guides specific substrates towards the Sec61 channel for insertion.

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Section: Sbh1 S3/t5 Phosphorylation Results In a Conformational Chang...mentioning

confidence: 76%

“…The intrinsically unstructured region of the Sec61β cytosolic domain can be crosslinked directly to ER-targeting sequences in the cytosolic vestibule of the mammalian Sec61 channel [20]. In the pathogenic yeast Cryptococcus neoformans , Sbh1 controls the biogenesis of secretory virulence factors [21].…”

Section: Introductionmentioning

confidence: 99%

The conserved membrane-proximal domain of Sbh1/ Sec61β guides signal peptides into the Sec61 channel

Yadhanapudi,

Bhadra,

Barbieri

et al. 2024

Preprint

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“…In vivo fungal virulence was tested in our murine inhalation model of cryptococcosis (12,(49)(50)(51).…”

Section: Murine Survival Studiesmentioning

confidence: 99%

“…A/J female mice (The Jackson Laboratories) were infected via the intranasal route as described above with 5 x 10 4 CFU of either WT (H99) or the pdr6Δ mutant strain. Mice were sacrificed as predetermined time points (14,35,50…”

Section: Histological Analysesmentioning

confidence: 99%

Lack of an atypical PDR transporter generates an immunogenicCryptococcus neoformansstrain that drives a dysregulated and lethal immune response in murine lungs

Winski,

Stuckey,

Marrufo

et al. 2024

Preprint

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Cryptococcus neoformansis an opportunistic fungal pathogen responsible for >150,000 deaths every year with a mortality rate as high as 81%. This high medical burden is due, in part, to an incomplete understanding of its pathogenesis. In a previous study, we identified a cryptococcal atypical pleiotropic drug resistance (PDR) transporter,PDR6, that regulated antifungal resistance and host interactions. Here, we follow-up on the role ofPDR6in cryptococcal virulence.In vivo, mice infected with thepdr6Δ strain display altered symptomatology and disease progression. Specifically, we observed a significant increase in the innate immune cell populations in thepdr6Δ-infected mice when compared to their WT-infected littermates. Furthermore, quantification of pulmonary cytokines/chemokines revealed a robust increase of pro-inflammatory cytokines in mice infected with thepdr6Δ mutant strain. Whereas antifungal treatment ofpdr6Δ-infected animals did not affect survival, treatment with a corticosteroid significantly extended survival, highlighting the importance of a balanced/controlled host immune response. We determined that the hyper-inflammatory immune response occurs, in part, because the loss of the Pdr6 transporter indirectly alters the cryptococcal cell wall architecture and results in the increased exposure of chitin, β-glucan, and other cryptococcal-specific pathogen associated molecular patterns. Taken together, this study provides clinical insights regarding cryptococcal pathogenesis while also providing additional functions of PDR-type ATP-binding cassette (ABC) transporters in pathogenic fungi.IMPORTANCEYeasts of theCryptococcusgenus, especiallyC. neoformans, can cause disease with unacceptably high mortality. This is due to delays in diagnostics, ineffective treatments, and an incomplete understanding of the interactions between this fungus and our immune system. In this study, we expand our knowledge of the biological function of thePDR6gene, particularly its effect on modulating the host’s immune response. Normally,C. neoformans’s infections are characterized by an anti-inflammatory response that is unable to control the yeast. In the absence ofPDR6, the response to the infection is a dysregulated pro-inflammatory response that initially controls the fungi but eventually results in death of the host due to too much tissue damage. This is due, in part, to an altered fungal surface. Given the dual role ofPDR6in modulating antifungal sensitivity and immune responses, this work provides important insights that may lead to new or improved therapeutics.

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The ER Protein Translocation Channel Subunit Sbh1 Controls Virulence of Cryptococcus neoformans

Cited by 2 publications

References 71 publications

The conserved membrane-proximal domain of Sbh1/ Sec61β guides signal peptides into the Sec61 channel

The conserved membrane-proximal domain of Sbh1/ Sec61β guides signal peptides into the Sec61 channel

Lack of an atypical PDR transporter generates an immunogenicCryptococcus neoformansstrain that drives a dysregulated and lethal immune response in murine lungs

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