Utility of DNA, RNA, Protein, and Functional Approaches to Solve Cryptic Immunodeficiencies

Margot A. Cousin, Matthew J. Smith, Ashley N. Sigafoos, Jay J. Jin, Marine I. Murphree, Nicole J. Boczek, Patrick R. Blackburn, Gavin R. Oliver, Ross A. Aleff, Karl J Clark, Eric D Wieben, Avni Y. Joshi, Pavel N. Pichurin, Roshini S. Abraham, Eric W Klee

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Purpose: We report a female infant identified by newborn screening for severe combined immunodeficiencies (NBS SCID) with T cell lymphopenia (TCL). The patient had persistently elevated alpha-fetoprotein (AFP) with IgA deficiency, and elevated IgM. Gene sequencing for a SCID panel was uninformative. We sought to determine the cause of the immunodeficiency in this infant. Methods: We performed whole-exome sequencing (WES) on the patient and parents to identify a genetic diagnosis. Based on the WES result, we developed a novel flow cytometric panel for rapid assessment of DNA repair defects using blood samples. We also performed whole transcriptome sequencing (WTS) on fibroblast RNA from the patient and father for abnormal transcript analysis. Results: WES revealed a pathogenic paternally inherited indel in ATM. We used the flow panel to assess several proteins in the DNA repair pathway in lymphocyte subsets. The patient had absent phosphorylation of ATM, resulting in absent or aberrant phosphorylation of downstream proteins, including γH2AX. However, ataxia-telangiectasia (AT) is an autosomal recessive condition, and the abnormal functional data did not correspond with a single ATM variant. WTS revealed in-frame reciprocal fusion transcripts involving ATM and SLC35F2 indicating a chromosome 11 inversion within 11q22.3, of maternal origin. Inversion breakpoints were identified within ATM intron 16 and SLC35F2 intron 7. Conclusions: We identified a novel ATM-breaking chromosome 11 inversion in trans with a pathogenic indel (compound heterozygote) resulting in non-functional ATM protein, consistent with a diagnosis of AT. Utilization of several molecular and functional assays allowed successful resolution of this case.

Original languageEnglish (US)
Pages (from-to)1-13
Number of pages13
JournalJournal of Clinical Immunology
DOIs
StateAccepted/In press - Apr 18 2018

Fingerprint

Exome
RNA
Ataxia Telangiectasia
Chromosomes, Human, Pair 11
DNA
Transcriptome
DNA Repair
Introns
Proteins
Ataxia Telangiectasia Mutated Proteins
Phosphorylation
IgA Deficiency
Neonatal Screening
Severe Combined Immunodeficiency
Lymphopenia
Lymphocyte Subsets
alpha-Fetoproteins
Heterozygote
Fathers
Immunoglobulin M

Keywords

  • ATM
  • DNA repair defects
  • flow cytometry
  • RNA sequencing
  • whole transcriptome sequencing
  • Whole-exome sequencing

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Cite this

Cousin, M. A., Smith, M. J., Sigafoos, A. N., Jin, J. J., Murphree, M. I., Boczek, N. J., ... Klee, E. W. (Accepted/In press). Utility of DNA, RNA, Protein, and Functional Approaches to Solve Cryptic Immunodeficiencies. Journal of Clinical Immunology, 1-13. https://doi.org/10.1007/s10875-018-0499-6

Utility of DNA, RNA, Protein, and Functional Approaches to Solve Cryptic Immunodeficiencies. / Cousin, Margot A.; Smith, Matthew J.; Sigafoos, Ashley N.; Jin, Jay J.; Murphree, Marine I.; Boczek, Nicole J.; Blackburn, Patrick R.; Oliver, Gavin R.; Aleff, Ross A.; Clark, Karl J; Wieben, Eric D; Joshi, Avni Y.; Pichurin, Pavel N.; Abraham, Roshini S.; Klee, Eric W.

In: Journal of Clinical Immunology, 18.04.2018, p. 1-13.

Research output: Contribution to journalArticle

Cousin, MA, Smith, MJ, Sigafoos, AN, Jin, JJ, Murphree, MI, Boczek, NJ, Blackburn, PR, Oliver, GR, Aleff, RA, Clark, KJ, Wieben, ED, Joshi, AY, Pichurin, PN, Abraham, RS & Klee, EW 2018, 'Utility of DNA, RNA, Protein, and Functional Approaches to Solve Cryptic Immunodeficiencies', Journal of Clinical Immunology, pp. 1-13. https://doi.org/10.1007/s10875-018-0499-6
Cousin, Margot A. ; Smith, Matthew J. ; Sigafoos, Ashley N. ; Jin, Jay J. ; Murphree, Marine I. ; Boczek, Nicole J. ; Blackburn, Patrick R. ; Oliver, Gavin R. ; Aleff, Ross A. ; Clark, Karl J ; Wieben, Eric D ; Joshi, Avni Y. ; Pichurin, Pavel N. ; Abraham, Roshini S. ; Klee, Eric W. / Utility of DNA, RNA, Protein, and Functional Approaches to Solve Cryptic Immunodeficiencies. In: Journal of Clinical Immunology. 2018 ; pp. 1-13.
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AU - Cousin, Margot A.

AU - Smith, Matthew J.

AU - Sigafoos, Ashley N.

AU - Jin, Jay J.

AU - Murphree, Marine I.

AU - Boczek, Nicole J.

AU - Blackburn, Patrick R.

AU - Oliver, Gavin R.

AU - Aleff, Ross A.

AU - Clark, Karl J

AU - Wieben, Eric D

AU - Joshi, Avni Y.

AU - Pichurin, Pavel N.

AU - Abraham, Roshini S.

AU - Klee, Eric W

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N2 - Purpose: We report a female infant identified by newborn screening for severe combined immunodeficiencies (NBS SCID) with T cell lymphopenia (TCL). The patient had persistently elevated alpha-fetoprotein (AFP) with IgA deficiency, and elevated IgM. Gene sequencing for a SCID panel was uninformative. We sought to determine the cause of the immunodeficiency in this infant. Methods: We performed whole-exome sequencing (WES) on the patient and parents to identify a genetic diagnosis. Based on the WES result, we developed a novel flow cytometric panel for rapid assessment of DNA repair defects using blood samples. We also performed whole transcriptome sequencing (WTS) on fibroblast RNA from the patient and father for abnormal transcript analysis. Results: WES revealed a pathogenic paternally inherited indel in ATM. We used the flow panel to assess several proteins in the DNA repair pathway in lymphocyte subsets. The patient had absent phosphorylation of ATM, resulting in absent or aberrant phosphorylation of downstream proteins, including γH2AX. However, ataxia-telangiectasia (AT) is an autosomal recessive condition, and the abnormal functional data did not correspond with a single ATM variant. WTS revealed in-frame reciprocal fusion transcripts involving ATM and SLC35F2 indicating a chromosome 11 inversion within 11q22.3, of maternal origin. Inversion breakpoints were identified within ATM intron 16 and SLC35F2 intron 7. Conclusions: We identified a novel ATM-breaking chromosome 11 inversion in trans with a pathogenic indel (compound heterozygote) resulting in non-functional ATM protein, consistent with a diagnosis of AT. Utilization of several molecular and functional assays allowed successful resolution of this case.

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KW - flow cytometry

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KW - whole transcriptome sequencing

KW - Whole-exome sequencing

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