Abstract
Molecular Misreading (MM) is the inaccurate conversion of genomic information into aberrant proteins. For example, when RNA polymerase II transcribes a GAGAG motif it synthesizes at low frequency RNA with a two-base deletion. If the deletion occurs in a coding region, translation will result in production of misframed proteins. During mammalian aging, misframed versions of human amyloid precursor protein (hApp) and ubiquitin (hUbb) accumulate in the aggregates characteristic of neurodegenerative diseases, suggesting dysfunctional degradation or clearance. Here cDNA clones encoding wild-type hUbb and the frame-shifted version hUbb(+1) were expressed in transgenic Drosophila using the doxycycline-regulated system. Misframed proteins were abundantly produced, both from the transgenes and from endogenous Drosophila ubiquitin-encoding genes, and their abundance increased during aging in whole-fly extracts. Over-expression of wild-type hUbb, but not hUbb(+1), was toxic during fly development. In contrast, when over-expressed specifically in adult flies, hUbb(+1) caused small decreases in life span, whereas hUbb was associated with small increases, preferentially in males. The data suggest that MM occurs in Drosophila and that the resultant misframed proteins accumulate with age. MM of the ubiquitin gene can produce alternative ubiquitin gene products with different and sometimes opposing phenotypic effects.
Highlights
The accurate read-out of genomic information into functional proteins is of critical importance to cellular homeostasis, and a significant disruption can lead to cell death [1, 2]
To determine if Molecular Misreading (MM) could be studied in Drosophila, cDNA clones encoding wild-type and frame-shifted versions of the human ubiquitin protein were expressed in Drosophila using the conditional doxycycline(DOX)regulated system (“Tet-on”) [55, 56]
DOX-dependent expression of human amyloid precursor protein (hApp)+1 protein was readily detected, using transgenes encoding hApp+1, as well as transgenes encoding wildtype hApp, and the hApp+1 protein became more abundant with age (Supplemental Figure S4), consistent with MM of the hApp construct
Summary
The accurate read-out of genomic information into functional proteins is of critical importance to cellular homeostasis, and a significant disruption can lead to cell death [1, 2]. It has been hypothesized that a loss of fidelity in information flow could contribute to aging through a feed-forward loop or “error catastrophe” in which errors lead to an increasing frequency of errors [3]. Aging in several cell types and species is associated with a progressive loss of nuclear genome integrity and structure that could potentially reduce fidelity of information flow from the nucleus [6,7,8]. Aging is characterized by significant changes in gene expression, including the tissue-specific induction of oxidative stress response genes and heat shock proteins (Hsps), and these gene expression changes may represent a response to mitochondrial malfunction, oxidative stress and proteotoxicity [9,10,11,12,13].
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