Brain transcriptome analysis reveals subtle effects on mitochondrial function and iron homeostasis of mutations in the SORL1 gene implicated in early onset familial Alzheimer’s disease

Abstract

Abstract

Background To prevent or delay the onset of Alzheimer’s disease (AD), we must understand its molecular basis. The great majority of AD cases arise sporadically with a late onset after 65 years of age (LOAD). However, rare familial cases of AD can occur due to dominant mutations in a small number of genes that cause an early onset prior to 65 years of age (EOfAD). As EOfAD and LOAD share similar pathologies and disease progression, analysis of EOfAD genetic models may give insight into both subtypes of AD. Sortilin-related receptor 1 (SORL1) is genetically associated with both EOfAD and LOAD and provides a unique opportunity to investigate the relationships between both forms of AD. Currently, the role of SORL1 mutations in AD pathogenesis is unclear.

Methods To understand the molecular consequences of SORL1 mutation, we performed targeted mutagenesis of the orthologous gene in zebrafish. We generated an EOfAD-like mutation, V1482Afs, and a putatively null mutation, to investigate whether EOfAD-like mutations in sorl1 display haploinsufficiency by acting through loss-of-function mechanisms. We performed mRNA-sequencing on whole brains comparing normal (wild type) fish with their siblings heterozygous for EOfAD-like or complete loss-of-function mutations in sorl1 or transheterozygous for these mutations. Differential gene expression and gene set enrichment analyses identified, respectively, changes in young adult zebrafish brain transcriptomes, and putative effects on neural subcellular functions.

Results We identified subtle effects on expression of genes involved in energy production, mRNA translation and mTORC1 signalling in both the EOfAD-like and null mutant brains, implying that these effects are due to sorl1 haploinsufficiency. Surprisingly, we also observed changes to expression of genes occurring only in the EOfAD-mutation carrier brains, suggesting gain-of-function effects. Transheterozygosity for the EOfAD-like and null mutations (i.e. lacking wild type sorl1), caused apparent effects on iron homeostasis and other transcriptome changes distinct from the single-mutation heterozygous fish.

Conclusions Our results provide insight into the possible early brain molecular effects of an EOfAD mutation in human SORL1. Differential effects of heterozygosity and complete loss of normal SORL1 expression are revealed.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

* Author full names and email addresses: Karissa Barthelson: karissa.barthelson{at}adelaide.edu.au, Stephen Martin Pederson: stephen.pederson{at}adelaide.edu.au, Morgan Newman: morgan.newman{at}adelaide.edu.au, Michael Lardelli: michael.lardelli{at}adelaide.edu.au

* Added supplemental data files

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Abbreviations

Aβ

amyloid beta

AD

Alzheimer’s disease

ANOVA

analysis of variance

APOE

apolipoprotein E

APP

amyloid beta a4 precursor protein

BDNF

brain-derived neurotrophic factor

bp

base-pair

Cas9

CRISPR associated protein 9

cDNA

complementary DNA

cox7a1

cytochrome c oxidase subunit 7A1

cpf1

CRISPR associated protein 12a

cpm

counts per million

CRISPR

clustered regularly interspaced short palindromic repeats

cuedc1b

cue domain containing 1b

DE

differentially expressed

DNaseI

deoxyribonuclease I

dqPCR

digital quantitative polymerase chain reaction

EGF

epidermal growth factor

EOfAD

early-onset familial Alzheimer’s disease

FC

fold change

FN

fibronectin

GEO

Gene Expression Omnibus

GSEA

gene set enrichment analysis

HIF1-α

hypoxia-inducible factor 1-alpha

hiPSCs

human induced pluripotent stem cells

ICD

intracellular domain

IRE

iron-responsive element

KEGG

Kyoto Encyclopedia of Genes and Genomes

LDLR

low density lipoprotein receptor

LOAD

late-onset Alzheimer’s disease

MAM

mitochondrial-associated membrane

mRNA

messenger RNA

MSigDB

molecular signatures database

mTORC

mammalian target of rapamycin complex

NMD

nonsense mediated mRNA-decay

nt

nucleotide

PAM

protospacer adjacent motif

PC

principal component

PCA

principal component analysis

PCR

polymerase chain reaction

PSEN

presenilin

RNA-seq

RNA sequencing

RT-PCRs

reverse transcription polymerase chain reactions

SAHMRI

South Australian Health and Medical Research Institute

siRNA

small interfering RNA

SORL1

sortilin-related receptor 1

TALEN

transcription activator-like effector nucleases

TCA

tricarboxylic acid

TMD

transmembrane domain

trans

transheterozygous

UTR

untranslated region

VPS10

vacuolar protein sorting 10

WT

wild type.