Saturation mutagenesis platform profiles stability effects of 2,696 variants across the globin gene family

A preprint posted to bioRxiv on 19 August 2026 reports the development and application of a modified Variant Abundance by Massively Parallel Sequencing (VAMP-seq) platform to the globin gene family. The researchers, using a lentiviral integration system in human cells, quantified the protein abundance effects of 2,696 amino acid substitutions in zeta-globin (HBZ), achieving near-complete coverage of possible single amino acid changes through saturation mutagenesis.

VAMP-seq — originally described in the context of PTEN and TPMT by Matreyek and colleagues — uses deep sequencing to quantify how each variant affects intracellular protein abundance as a proxy for protein stability and function. Variants that destabilise the protein lead to reduced abundance; those that are tolerated maintain near-wild-type levels.

The preprint integrates this experimental approach with a cross-paralog computational prediction framework, aiming to extrapolate stability effects from well-characterised globin family members — such as the extensively studied HBB (beta-globin) and HBA (alpha-globin) — to less-characterised paralogues. This is relevant because variants of uncertain significance (VUS) in embryonic and foetal globin genes are increasingly encountered in the context of expanded carrier screening and prenatal sequencing.

The authors argue that comprehensive functional maps of this kind — often called multiplexed assays of variant effect (MAVEs) — can inform variant classification under published frameworks such as those from the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. The work is methodologically relevant for researchers working on haemoglobinopathies, variant interpretation, and functional genomics.

This is a preprint and has not yet been peer-reviewed.