The operations within a living cell depend on the collective activity of networks of proteins, sometimes termed "interactomes". Within these networks, most proteins interact with few partners, while a small proportion of proteins, called hubs, participate in a large number of interactions and play a central role in organizing these interactomes. LC8 was first discovered as an essential component of the microtubule-based molecular motor dynein and as such is involved in fundamental processes, including retrograde vesicular trafficking, ciliary/flagellar motility, and cell division. More recently, evidence has accumulated that LC8 also interacts with proteins that are not clearly connected with dynein or microtubule-based transport, including some with roles in apoptosis, viral pathogenesis, enzyme regulation, and kidney development. Here, we introduce the idea that LC8 is a hub protein essential in diverse protein networks, and its function as a dynein light chain is but one of many. We further propose that the crucial regulatory roles of LC8 in various systems are due to its ability to promote dimerization of partially disordered proteins.

“What’s in a name? That which we call a rose By any other name would smell as sweet.” From “Romeo and Juliet”, William Shakespeare (1594) This article opens a series of publications on disambiguation of the basic terms used in the field of intrinsically disordered proteins. We start from the beginning, namely from the explanation of what the expression “intrinsically disordered protein” actually means and why this particular term has been chosen as the common denominator for this class of proteins characterized by broad structural, dynamic and functional characteristics.

Hydroxyproline (Hyp) O-glycosylation characterizes the hydroxyproline-rich glycoprotein (HRGP) superfamily of the plant extracellular matrix. Hyp glycosylation occurs in two modes: Arabinosylation adds short oligoarabinosides (Hyp-arabinosides) while galactosylation leads to the addition of larger arabinogalactan polysaccharides (Hyp-polysaccharides). We hypothesize that sequence-dependent glycosylation of small peptide motifs results in glycomodules. These small functional units in combination with other repetitive peptide modules define the properties of HRGPs. The Hyp contiguity hypothesis predicts arabinosylation of contiguous Hyp residues and galactosylation of clustered noncontiguous Hyp residues. To determine the minimum level of Hyp contiguity that directs arabinosylation, we designed a series of synthetic genes encoding repetitive (Ser-Pro 2 ) n , (Ser-Pro 3 ) n , and (Ser-Pro 4 ) n . A signal sequence targeted these endogenous substrates to the endoplasmic reticulum/Golgi for post-translational proline hydroxylation and glycosylation in transformed Nicotiana tabacum cells. The fusion glycoproteins also contained green fluorescence protein, facilitating their detection and isolation. The (Ser-Pro 2 ) n and (Ser-Hyp 4 ) n fusion glycoproteins yielded Hyp-arabinosides but no Hyp-polysaccharide. The motif (Ser-Pro 3 ) n was incompletely hydroxylated, yielding mixed contiguous/noncontiguous Hyp and a corresponding mixture of Hyparabinosides and Hyp-polysaccharides. These results plus circular dichroic spectra of the glycosylated and deglycosylated (Ser-Pro 2 ) n , (Ser-Pro 3 ) n , and (Ser-Pro 4 ) n modules corroborate the Hyp contiguity hypothesis and indicate that Hyp O-glycosylation is indeed sequence-driven. Hydroxyproline-rich glycoproteins (HRGPs)1 participate in the plant extracellular matrix as networks, exudates, and glycocalyx, comprising a superfamily that includes extensins (1), proline-rich proteins (PRPs) (2) and arabinogalactan-proteins (AGPs) (3). The three major families are distinguished by characteristic repetitive structural motifs: Ser-Hyp 4 in extensins, Pro-Hyp-Val-Tyr-Lys repeats and variants in PRPs, and XaaHyp-Xaa-Hyp repeats plus the presence of arabinogalactan polysaccharide in AGPs.The major post-translational modifications of HRGPs, proline hydroxylation and subsequent O-Hyp glycosylation, determine the properties of HRGPs to a greater or lesser extent. Carbohydrate accounts for as much as 95% of the hyperglycosylated AGPs and about 60% of extensins, thus forming the interactive molecular surface. In the lightly glycosylated PRPs, however, sugar may contribute as little as 1% of the mass.O-Hyp glycosylation occurs in two distinct modes, Hyp arabinosylation (4) and Hyp galactosylation (5), respectively. Hyp arabinosylation of virtually all HRGPs results in short (usually 1-4 residues), neutral, linear homooligosaccharides of L-arabinofuranose (Hyp-arabinosides). Hyp galactosylation, which is restricted to the AGPs, results in addition of much larger arabinogalactan heteropolysac...

The interactions of three subunits of cytoplasmic dynein from Drosophila melanogaster, LC8, Tctex-1, and the N-terminal domain of IC74 (N-IC74, residues 1-289), were characterized in vitro by affinity methods, limited proteolysis, and circular dichroism spectroscopy. These subunits were chosen for study because they are presumed to promote the assembly of the complex and to be engaged in the controlled binding and release of cargo. Limited proteolysis and mass spectrometry of N-IC74 in the presence of LC8 and Tctex-1 localized binding of Tctex-1 to the vicinity of K104 and K105, and localized binding of LC8 to the region downstream of K130. Circular dichroism, fluorescence, sedimentation velocity, and proteolysis studies indicate that N-IC74 has limited secondary and tertiary structure at near physiological solution conditions. Upon addition of LC8, N-IC74 undergoes a significant conformational change from largely unfolded to a more ordered structure. This conformational change is reflected in increased global protection of N-IC74 from proteolytic digestion following the interaction, and in a significant change in the CD signal. A smaller but reproducible change in the CD spectra was observed upon Tctex-1 binding as well. The increased structure introduced into N-IC74 upon light chain binding suggests a mechanism by which LC8 and Tctex-1 may regulate the assembly of the dynein complex.