AstraZeneca and Lieber Institute researchers use RNA-Seq to discover novel protein associated with schizophrenia

Study results indicate that discovery of recently evolved protein could lead to new targets for neuropsychiatric disorders and novel treatments for patients

AstraZeneca and the Lieber Institute for Brain Development (LIBD), an affiliate of the Johns Hopkins University School of Medicine, first partnered in 2013. Ever since, scientists from both partners have worked side-by-side to interrogate emerging genetic data for neuropsychiatric and neurodevelopmental disorders, utilising genomics and induced pluripotent stem cell based models. The joint goal is to define new approaches to treating diseases of the brain by combining the expertise and experience of both partners.

The team’s latest study results were published in the journal Nature Medicine. This is the first study to identify a human-specific form of a protein related to AS3MT that represents new insight into the causative mechanisms of schizophrenia and related disorders, including depression, bipolar disorder and attention deficit hyperactivity disorder.

The publication describes how the team went beyond previously reported research to understand the molecular drivers for the association between the chromosomal region 10q24.32 and neuropsychiatric disorders.  By performing deep RNA sequencing on post-mortem human brain samples they identified over-expression of a novel human isoform of AS3MT named ASMTd2d3 as the genetic risk-factor.

This pioneering discovery has enabled the team to progress into pluripotent stem cell models where they can begin to understand how ASMTd2d3 effects developing and mature human neurons.  These efforts may identify future molecular targets for which we can design novel therapeutics to reverse the damage done by ASMTd2d3.

Characterizations of AS3MT isoforms

rna-seq

(a) Primary AS3MT transcripts in human brain. Black rectangles are translated regions, and white rectangles are untranslated regions. The novel junction E1–E4 is marked in red; the junction E2–E3 is marked in orange. (b) RNA-seq analysis of AS3MTd2d3 isoform in diverse human tissues from the GTEx data set (total n = 8,193); x axis represents log2(RPM) of AS3MTd2d3 isoform. Error bars represent mean ± s.d. ***P = 2.35 × 10−34. The P value of fold-change difference is calculated using linear mixed-effects regression modeling. The data in “brain” (red bar) is the average of diverse brain tissues (black bars), whereas data in “somatic tissues” (blue bar) is the average of nonbrain tissues (c) Representative western blots of AS3MTd2d3 and AS3MTfull isoforms from pools of multiple wells (n > 3) of transfected HEK293 and B104 cells. Each experiment was replicated at least three times. GAPDH, glyceraldehyde 3-phosphate dehydrogenase. (d) Representative protein expression of AS3MTd2d3 and AS3MTfull in commercial human brain extracts (pooled from n > 10 brains). Positive control lanes include HEK293 cells transfected with AS3MTd2d3 and AS3MTfull. Each experiment was replicated at least three times. (e) Representative subcellular localization of AS3MTd2d3 (left) and AS3MTfull (right) proteins in B104 cells. Cells expressing Flag-tagged AS3MT isoforms were stained with an anti-Flag antibody to determine subcellular protein distribution (red). Expression of AS3MT does not co-localize with the nuclear counter stain DAPI (blue). Visualization of approximately 20–30 cells across separate cultures was used for assessment, and representative transfected cells are shown. Each experiment was replicated at least three times. Scale bars, 20 μm. (f) Association of rs7085104 with AS3MTd2d3 (left, P = 5.0 × 10−13) and AS3MTfull (right, P = 0.627) expression in the Caucasian DLPFC RNA-seq samples (n = 322). P values were calculated by linear regression using log2(RPM + 1). Error bars depict mean ± s.d. (g) Association of rs7085104 with AS3MTd2d3 (left, P = 8.1 × 10−3) and AS3MTfull (right, P = 0.329) expression in the frontal cortex samples from GTEx data set (n = 99). P values were calculated by linear regression using log2(RPM + 1). Error bars depict mean ± s.d.

The findings were published in the May 2016 issue of Nature Medicine.

Source – AstraZeneca

Li M, Jaffe AE, Straub RE, Tao R, Shin JH, Wang Y, Chen Q, Li C, Jia Y, Ohi K, Maher BJ, Brandon NJ, Cross A, Chenoweth JG, Hoeppner DJ, Wei H, Hyde TM, McKay R, Kleinman JE, Weinberger DR. (2016) A human-specific AS3MT isoform and BORCS7 are molecular risk factors in the 10q24.32 schizophrenia-associated locus. Nat Med [Epub ahead of print]. [abstract]

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