# Automated Forward Genetic Analysis of Adaptive Immunity

> **NIH NIH R01** · UT SOUTHWESTERN MEDICAL CENTER · 2020 · $1,617,654

## Abstract

PROJECT SUMMARY
To understand immunity mechanistically, one must identify its constituent parts: those proteins with non-
redundant function in the exercise of any immune process, however we choose to define it. One might assume
that most of the essential proteins are known, but they are not. This conclusion is based on growing
experience with a new method in mammalian genetics, developed in our laboratory1. By damaging or
destroying genes at random with ENU, sequencing the whole exome of every G1 male carrier of these
mutations, and pre-genotyping all G3 animals at potential mutation sites prior to phenotypic screening, we are
able to determine which mutations cause phenotype in real time. In effect, we now positionally clone
instantaneously: when a phenotype is seen, its cause is known. We also know precisely which genes we have
altered, what change was made, and how many times the mutation was tested for phenotypic effects in the
homozygous state. Over the 20 months prior to submission of this proposal, we applied this method to 30,446
G3 mutant mice from 1,189 pedigrees. Each mutant mouse was subjected to 110 immunological assays
testing the immunological effects of 70,144 coding or splicing changes in 17,640 genes. For 1,865 genes, at
least one putative null allele was examined three or more times in the homozygous state. We estimate that we
have stringently tested 18.4% of all genes in the genome for a necessary role in those immune functions falling
within our sphere of interest. In so doing, we have identified many proteins essential for one immunological
process or another, some of them known, but most of them previously unrecognized1. This preliminary survey
makes it clear that much remains to be learned about phenomena one might regard as well studied. What are
the requirements for a T-dependent antibody response? We may think we know, but in fact, many of the
essential proteins are still undiscovered. As more than 100 new components of the immune system have been
identified through mutagenesis, it is incumbent upon us to understand how at least some of the mutations
produce their phenotypic effects. We have chosen to address a subset of genes, within which mutations cause
striking phenotypes bearing on adaptive immunity. First, we will probe the basis of allergic responses by
studying a set of four genes in which mutations cause exaggerated IgE responses to injected papain and/or
ovalbumin/alum immunization. Second, we will examine mutations in three genes not generally known to be
associated with immunity that dramatically affect lymphocyte development and/or function. Third, we will study
the regulation and function of IgD in light of a mutation that causes a selective hyper-IgD syndrome. We will
also continue to mutagenize and screen mice for still other immune defects, and make our data accessible to
the scientific community via software developed for this purpose.

## Key facts

- **NIH application ID:** 9929531
- **Project number:** 5R01AI125581-05
- **Recipient organization:** UT SOUTHWESTERN MEDICAL CENTER
- **Principal Investigator:** BRUCE A BEUTLER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $1,617,654
- **Award type:** 5
- **Project period:** 2016-06-20 → 2021-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9929531

## Citation

> US National Institutes of Health, RePORTER application 9929531, Automated Forward Genetic Analysis of Adaptive Immunity (5R01AI125581-05). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/9929531. Licensed CC0.

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