# Exploratory Studies of SPPL3

> **NIH NIH R21** · JOHNS HOPKINS UNIVERSITY · 2020 · $204,688

## Abstract

PROJECT SUMMARY/ABSTRACT
A variety of important biological pathways in health and disease are regulated by intramembrane proteolysis,
the process that achieves the targeted cleavage of protein substrates within or proximal to the lipid bilayer.
Four classes of intramembrane proteases have been described, including the site 2 protease (S2P)
metalloprotease family, the rhomboid serine protease family, the Rce1 glutamyl protease family, and the
aspartyl protease family that includes presenilins and the related Signal Peptide Peptidase (SPP)-like (SPPL)
subfamily. The aspartyl proteases of the SPPL subfamily, including SPP, SPPL2a, SPPL2b, SPPL2c, and
SPPL3, are perhaps the least understood subset of intramembrane proteases. Only recently have the
biological roles of some of these proteases emerged, and only a handful of relevant biological substrates have
been identified. How these intramembrane proteases specifically recognize and cleave their substrates
remains mysterious. Recently, we discovered a critical role for SPPL3 in NK cell maturation and cytotoxicity.
SPPL3 is required in a cell-autonomous manner for the maturation of NK cells from the immature
CD27+CD11b- stage to the CD27+CD11b+ and CD27-CD22b+ stages, and for normal NK cell cytotoxicity
toward tumor cell targets. Mice engineered to express only SPPL3 D271A in the NK lineage revealed that the
proteolytic function of SPPL3 is required in NK cell maturation and function. Like other aspartyl intramembrane
proteases of the SPPL subfamily, very little is known about how SPPL3 recognizes and cleaves its substrates.
All SPPL proteases possess YD and GXGD motifs that contain the catalytic aspartates, but other regions in the
protein that are required for substrate recognition and cleavage have not been identified. It is also unclear
whether a specific amino acid sequence in a putative substrate is recognized by SPPL3 or whether other
features of the target, such as the size or composition of cytoplasmic and luminal domains, facilitate
recognition by SPPL3. Furthermore, although some SPPL3 substrates have been identified, the relevant
substrate that must be cleaved by SPPL3 during NK cell maturation is currently unknown. To expand our basic
understanding of SPPL3 and related aspartyl intramembrane proteases, and to advance understanding of the
key checkpoint in NK cell maturation controlled by SPPL3, we will 1) use a novel high-throughput assay for
SPPL3 protease activity to identify residues required for substrate recognition and cleavage; 2) determine
whether substrate binding and cleavage are controlled by different SPPL3 residues, and whether the same
SPPL3 determinants are required for the cleavage of four biochemically distinct substrates; 3) address what
features in a substrate allow it to be recognized and cleaved by SPPL3; and 4) identify SPPL3 substrates in
NK cells. Our results will expand understanding of intramembrane aspartyl proteases and illuminate molecular
events that control ...

## Key facts

- **NIH application ID:** 9822180
- **Project number:** 5R21AI143053-02
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Joel L Pomerantz
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $204,688
- **Award type:** 5
- **Project period:** 2018-11-15 → 2020-10-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9822180

## Citation

> US National Institutes of Health, RePORTER application 9822180, Exploratory Studies of SPPL3 (5R21AI143053-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9822180. Licensed CC0.

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