# Membrane protein localization and function during ciliary signaling and cell-cell fusion

> **NIH NIH R35** · UNIV OF MARYLAND, COLLEGE PARK · 2021 · $547,200

## Abstract

We will use fertilization in the bi-ciliated green alga Chlamydomonas as a model system to investigate
conserved cellular and molecular mechanisms of ciliary signaling and the gamete membrane fusion reaction.
When Chlamydomonas gametes of opposite types are mixed together they adhere to each other by
complementary adhesion receptors on their cilia. Interaction between the adhesion receptors (encoded by
SAG1 in plus gametes and SAD1 in minus gametes) initiates a signaling pathway within the cilia that is
transmitted to the cell body and 1) triggers the gametes rapidly (~ 5 minutes) to undergo a striking
redistribution of pre-existing ciliary adhesion polypeptides from the cell body plasma membrane to the base of
the cilium, followed by trafficking onto the ciliary membrane; and 2) uncovers and activates apically localized
protrusions - - the plus and minus mating structures - - on the cell body of each gamete between the two cilia.
Each mating structure bears a gamete-specific adhesion protein, distinct from those on the cilia, that bind the
tips of the two mating structures together. Mating structure adhesion is rapidly followed by lipid bilayer
merger through the action of the gamete-specific broadly conserved protein, HAP2. Soon after fusion, the
mating structure adhesion molecules and the ciliary adhesion receptors receptors are down-regulated. In our
ciliary signaling and protein trafficking studies, we have found that ciliary adhesion-induced apical
localization of SAG1 polypeptide depends on singlet microtubules in the cytoplasm and action of the
retrograde intraflagellar transport (IFT) motor. On the other hand, contrary to current models, dynamic
trafficking of SAG1 into cilia does not require the anterograde IFT motor. Finally, we determined that SAG1
movement into cilia is uni-directional. During down-regulation, SAG1 does not return to the cell body, but the
entire pre-existing complement of the protein is shed into the medium in the form of ciliary ectosomes. Our
findings challenge existing, largely untested models of ciliary membrane protein trafficking and set the stage
for new strategies to investigate cellular and molecular mechanisms of the dynamic regulation of the protein
composition of cilia. In our studies of the gamete membrane fusion reaction, we have identified the missing
member of a receptor pair essential for adhesion between the plus and minus mating structures. And, in what
we feel is a major advance in the field, we discovered that the fusion-essential HAP2 protein is a Class II fusion
protein in the same family as Dengue and Zika virus fusion proteins. Mutational or immunological
interference with the HAP2 “fusion loop” does not interfere with HAP2 localization to the mating structure,
but renders HAP2 inactive in bilayer fusion. Our discoveries make possible a detailed, structure- and function-
based molecular dissection of eukaryotic gamete fusion. Furthermore they open the possibility that strategies
used to bloc...

## Key facts

- **NIH application ID:** 10152601
- **Project number:** 5R35GM122565-05
- **Recipient organization:** UNIV OF MARYLAND, COLLEGE PARK
- **Principal Investigator:** William J Snell
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $547,200
- **Award type:** 5
- **Project period:** 2017-05-01 → 2022-09-16

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10152601

## Citation

> US National Institutes of Health, RePORTER application 10152601, Membrane protein localization and function during ciliary signaling and cell-cell fusion (5R35GM122565-05). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10152601. Licensed CC0.

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