The unresolved puzzle why alanine extensions cause disease

Abstract

The prospective increase in life expectancy will be accompanied by a rise in the number of elderly people who suffer from ill health caused by old age. Many diseases caused by aging are protein misfolding diseases. The molecular mechanisms underlying these disorders receive constant scientific interest. In addition to old age, mutations also cause congenital protein misfolding disorders. Chorea Huntington, one of the most well-known examples, is caused by triplet extensions that can lead to more than 100 glutamines in the N-terminal region of huntingtin, accompanied by huntingtin aggregation. So far, nine disease-associated triplet extensions have also been described for alanine codons. The extensions lead primarily to skeletal malformations. Eight of these proteins represent transcription factors, while the nuclear poly-adenylate binding protein 1, PABPN1, is an RNA binding protein. Additional alanines in PABPN1 lead to the disease oculopharyngeal muscular dystrophy (OPMD). The alanine extension affects the N-terminal domain of the protein, which has been shown to lack tertiary contacts. Biochemical analyses of the N-terminal domain revealed an alanine-dependent fibril formation. However, fibril formation of full-length protein did not recapitulate the findings of the N-terminal domain. Fibril formation of intact PABPN1 was independent of the alanine segment, and the fibrils displayed biochemical properties that were completely different from those of the N-terminal domain. Although intranuclear inclusions have been shown to represent the histochemical hallmark of OPMD, their role in pathogenesis is currently unclear. Several cell culture and animal models have been generated to study the molecular processes involved in OPMD. These studies revealed a number of promising future therapeutic strategies that could one day improve the quality of life for the patients.

Methylation of the nuclear poly(A)-binding protein by type I protein arginine methyltransferases – how and why

“…For example, the self-association of the yeast protein Npl3p is also increased in the absence of arginine methylation (Yu et al , 2004 ;McBride et al , 2007 ), and we have noticed a strongly increased background in immunoprecipitations from PRMT1 -/-cells, consistent with increased protein aggregation in the absence of methylation (Ostareck -Lederer et al, 2006 ). As the formation of fibrillar aggregates of PABPN1 is important in the etiology of OPMD (Brais et al , 1998 ;Calado et al , 2000b ;Winter et al , 2013 ), an effect of methylation on the aggregation of the protein would be of particular interest. However, although several assays demonstrated a higher tendency of recombinant PABPN1 purified from E. coli to oligomerize or aggregate compared with the fully methylated protein purified from calf thymus, this was traced to the presence of an N-terminal His-tag, not the lack of methylation: the recombinant protein lacking the tag showed no more self-association than the methylated protein from calf thymus in pull-down experiments and did not display any aggregation in gel-shift experiments for binding to poly(A) (Fronz et al , 2008 ).…”

“…PABPC, which is important in translation and the control of RNA stability, is composed of four RNA-binding domains of the RNA recognition motif (RRM) or ribonucleoprotein (RNP) type, followed by a linker and a conserved C-terminal α -helical domain. Although the nuclear poly(A)-binding protein (PABPN1 in humans) shares the RNA binding specificity of PABPC, its structure is quite different (Nemeth et al , 1995 ) (Figure 2 ): in the order N-terminus to C-terminus, it is composed of a polyalanine sequence, expansions of which cause the human genetic disease oculopharyngeal muscular dystrophy (OPMD) (Brais et al , 1998 ;Calado et al , 2000b ;Winter et al , 2013 ); an unstructured acidic domain of unknown function; a coiled coil domain, predicted on the basis of its characteristic pattern of hydrophobic amino acid side chains and confirmed by CD spectroscopy (Kerwitz et al , 2003 ); a single central RRM domain; finally, an unstructured basic C-terminal domain. The C-terminal domain is 49 amino acids long and contains 13 arginine residues, all of which are asymmetrically dimethylated, as determined by a combination of mass spectrometry and Edman sequencing (Smith et al , 1999 ).…”

“…However, at a concentration of ∼ 2 μ m , the mammalian PABPN1 is predominantly monomeric, as determined by analytical ultracentrifugation (Nemeth et al , 1995 ;Meyer et al , 2002 ;Kerwitz et al , 2003 ). Various types of aggregation require higher protein concentrations (Winter et al , 2013 ). The RNA-binding unit is also a monomer (Meyer et al , 2002 ;Song et al , 2008 ).…”

Influence of the polypeptide environment next to amyloidogenic peptides on fibril formation

“…It is not clear whether an alanine extension-induced misfolding of the respective protein constitutes the prime pathogenesis factor (reviewed by Winter et al, 2013). Exclusively nucleic binding proteins have so far been identified to be associated with disease when alanine segments are expanded.…”

“…Alanine peptides have been called conformational chameleons because they display the whole spectrum, from high to low α-helicities, depending on the flanking residues (Miller et al, 2001). Despite the high propensity of alanine segments to form α-helical structures, they also show a strong tendency to fibrillize (Forood et al, 1995;Nguyen and Hall, 2005;Winter et al, 2013). The high stability of fibrils from alanine containing segments or poly-alanines raises the question: what thermodynamic parameters would lead to fibril formation of this type?…”