# The roles of ceramide and its derivatives in A. phagocytophilum pathogenesis

> **NIH NIH R01** · VIRGINIA COMMONWEALTH UNIVERSITY · 2024 · $700,747

## Abstract

Anaplasma phagocytophilum (Ap) is an obligate intracellular bacterium that causes the emerging and potentially
fatal infection, human granulocytic anaplasmosis (HGA). The microbial-host interactions that facilitate Ap
intracellular proliferation and dissemination have remained poorly characterized. We uncovered a novel role for
the host bioactive sphingolipid, ceramide-1-phosphate (C1P), as a regulator of Golgi fragmentation. Specifically,
C1P activates a PKC/Cdc42/JNK signaling axis that phosphorylates GRASP55 (Golgi reassembly stacking
protein 55 kDa) to destabilize the trans-Golgi network (TGN) and amplify TGN anterograde traffic. We determined
that Ap infection upregulates C1P synthesis and induces Golgi fragmentation in a C1P signaling-dependent
manner. Multivesicular bodies (MVBs) are endosomal sorting stations that receive Golgi traffic. MVB limiting
membrane invagination and scission deliver cargo into the MVB in intraluminal vesicles (ILVs). ILV membranes
are enriched in cholesterol, sphingolipids, and sphingolipid metabolic enzymes that promote cholesterol release
from ILVs. MVB fusion with the plasma membrane releases ILVs as exosomes, which induce a battery of
responses in recipient cells including signaling, immunity, and inflammation. The importance of exosome-
mediated cell-to-cell communication in infectious disease is just beginning to be appreciated. We discovered that
Ap resides in a pathogen-modified MVB that receives sphingomyelin-rich TGN cargo as ILVs. The bacterium
coopts MVB lipid metabolic machinery to parasitize sphingolipids and cholesterol, which is essential for its growth
and conversion from its replicative to infectious form. Consistent with MVB exosome release, the Ap vacuole
fuses with the plasma membrane in a Rab27a-dependent manner to disperse infectious progeny to naïve cells.
We also found that Ap alters exosome content. In this competitive renewal, we will interrogate our hypothesis
that Ap induces C1P to destabilize the Golgi and promote TGN-to-ApV trafficking of SM-rich vesicles that enables
Ap to parasitize host lipids for proliferation and dissemination. We will also test our hypothesis that Ap modulates
the proteomic and lipidomic content of ILVs that, when released as exosomes together with progeny bacteria,
synergistically benefit infection of naïve cells and/or contributes to immunopathology associated with HGA.
Completing the Aims herein will yield one of the most refined models for intracellular bacterial pathogenesis,
further illuminate our newly discovered roles of C1P as both a regulator of Golgi stability and microbial target for
host modulation, and further understanding of how exosomes contribute to pathogenic processes.

## Key facts

- **NIH application ID:** 10861876
- **Project number:** 5R01AI139072-07
- **Recipient organization:** VIRGINIA COMMONWEALTH UNIVERSITY
- **Principal Investigator:** CHARLES E. CHALFANT
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $700,747
- **Award type:** 5
- **Project period:** 2018-06-06 → 2028-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10861876

## Citation

> US National Institutes of Health, RePORTER application 10861876, The roles of ceramide and its derivatives in A. phagocytophilum pathogenesis (5R01AI139072-07). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10861876. Licensed CC0.

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