Functional Domains
Two complementary domain definitions — sequence-based (Pfam) and structure-based (TED) — mapped onto splicing events without prejudging the outcome.
SPLISOFORMS scans every isoform's protein sequence against the full Pfam-A HMM library with HMMER (via pyhmmer), recording every significant domain hit with residue-level coordinates. Each hit is then annotated along two independent axes:
- Functional class — what the domain does, based on a curated Pfam clan + accession mapping.
- Alteration type — where the splicing event has perturbed the domain (N-terminal, C-terminal, both, internal, or full loss / gain).
The Mapping Pipeline
The key idea is that an isoform and its canonical reference go through the same scan — so any difference between them reflects the splice event, not a change in method. Mapping runs in three steps:
- Scan. Each isoform's protein sequence is searched against Pfam-A using Pfam's own gathering (GA) score thresholds. Every hit is recorded with its residue coordinates and how much of the domain model it covers, so a partial hit can be pinned to a specific part of the domain.
- Compare. The same scan is run on the gene's canonical isoform. Matching each hit to the canonical hit for the same Pfam family — and comparing how much of the model each one covers — is what determines the alteration type.
- Classify. Every hit is also tagged with a functional class derived from its Pfam clan and accession, describing what the domain does.
Functional Classes
The classes below carve up the Pfam universe into broad activity buckets that surface together in the UI and the API. The taxonomy is intentionally coarse and conservative — curated assignments only. Domains whose Pfam family has no clan assignment, or whose clan we haven't reviewed yet, land in Other / unclassified. That bucket is not "low importance" — it's unsorted, and many transcription co-factors, RNA-processing factors, and structural adaptors live there. Treat the class as a filter / facet, not a verdict.
| Class | Family | Example Pfam IDs |
|---|---|---|
| Kinase | catalytic | Pkinase (PF00069), Pkinase_Tyr (PF07714) |
| Phosphatase | catalytic | Y_phosphatase (PF00102), DSPc (PF00782), PP2C (PF00481) |
| Protease | catalytic | Trypsin (PF00089), Peptidase_C1 (PF00112), Caspase (PF00656) |
| Catalytic (other) | catalytic | DEAD helicase (PF00270), Rossmann-fold dehydrogenases, P-loop NTPases |
| Ubiquitin ligase / E2 | catalytic | RING (PF00097), HECT (PF00632), UQ_con / E2 (PF00179) |
| DNA-binding | binding | Homeobox (PF00046), HLH (PF00010), bZIP (PF00170), C2H2 zinc finger (PF00096) |
| RNA-binding | binding | RRM (PF00076), KH (PF00013), dsrm (PF00035) |
| Signaling adapter | adapter | SH2 (PF00017), SH3 (PF00018), PDZ (PF00595), PH (PF00169), WW (PF00397) |
| Oligomerization / scaffold | scaffold | Ankyrin (PF00023), WD40 (PF00400), TPR clan (CL0020) |
| Other / unclassified | other | Anything outside the curated list — explicitly NOT a low-priority bucket; many TFs and RNA-processing factors fall through here when their Pfam family lacks a clan assignment. |
Alteration Geometry
Each domain hit carries an alteration_type describing where the splicing event has changed the domain relative to the canonical isoform's hit on the same Pfam family. This matters because the same family can be activated, deactivated, or converted into a dominant-negative version depending on which piece of the model is removed.
Full-coverage HMM hit (>= 80% of the model length). The domain looks complete in this isoform.
The hit starts well into the domain model, so the N-terminal part of the canonical domain is missing. Depending on the domain this can strip an auto-inhibitory region or remove catalytic machinery (e.g. the kinase N-lobe) — the effect depends on exactly which residues are gone.
The hit ends well before the domain model does, so the C-terminal part is missing. This can drop regulatory tails, docking sites or substrate-recognition loops while sparing the catalytic core — or remove the core itself.
An internal slice of the HMM model is retained while both ends are missing. Often the result of an exon-skipping event that lands inside the domain.
Both ends are inside the HMM model and HMM coverage is below 50% — consistent with a small fragment surviving inside an otherwise lost domain, or an internal in-frame deletion.
The domain is present in the canonical isoform but entirely absent from the novel one. Shown only on the canonical reference track; absent from the isoform track.
The domain matches in the novel isoform but not the canonical — usually a novel exon or a frameshift that exposes a different reading frame.
How the cut-offs work
A domain counts as intact when the isoform covers at least 80% of the domain model (or at least 85% of what the canonical isoform covers for the same family). Whether a truncation is called N- or C-terminal depends on which end is missing: a hit that only starts after the first 10% of the model has lost its N-terminus, and one that ends before the last 10% has lost its C-terminus.
TED — Structural Domains
While Pfam answers "which sequence motifs are present", TED (The Encyclopedia of Domains) answers the complementary, structure-based question: how does the folded protein partition into independent, compact domain units?
Uniform de novo domain chopping
Because every isoform has its own AlphaFold 3 structure, canonical and novel isoforms are both segmented de novo with Chainsaw. Every isoform is therefore evaluated on the same footing, with domain boundaries drawn purely from its 3D fold rather than inherited from the reference.
Why two systems?
A splice event can leave a Pfam motif intact while collapsing the structural domain that hosts it (or vice-versa). Reading the two side by side separates "the sequence signature survives" from "the fold survives".