Truncated variants of hyaluronan-binding protein 1 bind hyaluronan and induce identical morphological aberrations in COS-1 cells.
Hyaluronan (HA)-binding protein 1 (HABP1) is multifunctional in nature and exists as a trimer through coiled-coil interaction between alpha-helices at its N- and C-termini. To investigate the importance of trimeric assemblage and HA-binding ability of HABP1, we generated and overexpressed variants of HABP1 by truncating the alpha-helices at its termini. Subsequently, these variants were transiently expressed in COS-1 cells to examine the influence of these structural variations on normal cell morphology, as compared with those imparted by HABP1. Substantiating the centrality of coiled-coil interaction for maintaining the trimeric assembly of HABP1, we demonstrate that disruption of trimerization does not alter the affinity of variants towards its ligand HA. Transient expression of HABP1 altered the morphology of COS-1 cells by generating numerous cytoplasmic vacuoles along with disruption of the f-actin network. Interestingly, the truncated variants also imparted identical morphological changes. Characterization of the cytoplasmic vacuoles revealed that most of these vacuoles were autophagic in nature, resembling those generated under stress conditions. The identical morphological changes manifested in COS-1 cells on transient expression of HABP1 or its variants is attributed to their comparable HA-binding ability, which in concert with endogenous HABP1, may deplete the cellular HA pool. Such quenching of HA below a threshold level in the cellular milieu could generate a stress condition, manifested through cytoplasmic vacuoles and a disassembly of the f-actin network.
Sengupta
Aniruddha
A
Biochemistry Laboratory, School of Environmental Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
Tyagi
Rakesh K
RK
Datta
Kasturi
K
D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D019556:COS Cells / Q000737:chemistry / Q000378:metabolism / Q000473:pathology* / Q000821:virology; D002461:Cell Line, Transformed; D002522:Chlorocebus aethiops; D003599:Cytoskeleton / Q000235:genetics / Q000473:pathology; D014644:Genetic Variation / Q000235:genetics*; D018960:Hyaluronan Receptors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D006820:Hyaluronic Acid / Q000378:metabolism*; D011485:Protein Binding / Q000235:genetics; D013539:Simian virus 40 / Q000235:genetics
D018960:Hyaluronan Receptors; D006820:Hyaluronic Acid