Calcium-binding proteins in the vorticellid spasmoneme

Abstract

The proteins of the contractile spasmoneme from Vorticella convallaria, Carchesium polypinum, and Zoothamnium geniculatum have been extracted in the detergent, sodium dodecyl sulfate (SDS), as well as urea and guanidine hydrochloride (GuCl). After SDS extraction, the molecular weight distribution of the proteins was examined by means of SDS-polyacrylamide gel electrophoresis. Significant amounts of material corresponding to the contractile proteins actin and tubulin are not present.The contractile organelles in the three species examined contain a group of closely related proteins of molecular weight near 20,000, which constitute a major part (40-60%) of the dry mass. The 20,000 mol wt proteins in Zoothamnium bind calcium with high affinity (pK -~ 6) and are termed "spasmins." By means of urea polyacrylamide gel electrophoresis, it is demonstrated that in Carchesium and Zoothamnium certain spasmin components bind calcium even in the presence of 6 M urea. The binding of calcium in 6 M urea suggests a functional relationship between the spasmins and the calcium-binding proteins of striated muscle which behave similarly. The calcium binding in urea also indicates that the spasmins within a single spasmoneme have different calcium affinities, and this difference in calcium-binding properties may be an important factor in the physiological function of the organelle. KEY WORDS contraction calcium Vorticella spasmonemeSeveral species of ciliated protozoa exhibit a rapid form of contraction. This is particularly clear in the stalked peritrichous ciliates, Vorticella, Carchesium, and Zoothamnium, which, on contraction, throw their stalks into tight helical coils or folds. Within the stalk is the contractile organelle, often referred to as the myoneme (1), though the term spasmoneme (2) is preferable since recent studies indicate that the organelle is biochemically distinct from the myosin-and actin-based contractile systems (3).The spasmoneme contracts within 2-10 ms (4, 5), and the shortening velocity may be as high as 172 lengths/s, whereas the extension phase is of the order of seconds. The contraction velocity of the spasmoneme in Zoothamnium, (measured in lengths/second), is thus nine times greater than in 358J. CELL BIOLOGY 9 The Rockefeller University Press 9

Cloning, Localization, and Axonemal Function ofTetrahymenaCentrin

“…Centrin is an integral part of microtubule-organizing centers, such as centrioles and basal bodies, and of the filamentous structures associated with these regions (Salisbury et al, 1984) including flagellar roots. Centrin is also part of the family of proteins that make up contractile stalks and fibers in protozoa such as Stentor or Vorticella (Routledge, 1978;Maciejewski et al, 1999). It may be part of the microtubule-severing apparatus at the base of the cilium (Sanders and Salisbury, 1989) and, most significantly for this study, it is found as a part of some of the inner dynein arms of Chlamydomonas axonemes (LeDizet and Piperno, 1995).…”

Mechanics of Vorticella Contraction

“…The only calcium-binding proteins in the spasmoneme are the spasmins (12,13), which are integral to the spasmonemal structure (20). Hence, there are practically no calcium-binding proteins in the aqueous phase inside the spasmoneme.…”

“…The released Ca 2þ ions bind to a 20-kDa calcium-binding protein called spasmin, which constitutes 40-60% of the spasmoneme dry mass (12,13). As a result, a state of tension is induced in the spasmoneme which drives its contraction with a maximum speed of~6 cm s À1 (Fig.…”

Identification and Localization of a Protein Immunologically Related to Caltractin (Centrin) in the Myonemes and Membranelles of the Heterotrich Ciliate Stentor coeruleus

“…It has been assumed for some time that the myonemal filaments of the heterotrichs were likely to be composed of spasmin, a protein first isolated from spasmonemes of Zoothamnion and Carchesium (Amos et al 1975; Routledge 1978) and recently cloned in Vorticella (Maciejewski et al 1999). Spasmin, like caltractin, is an acidic protein with an MW of 20 kDa that has an altered electrophoretic mobility in the presence of Ca 2+ , but it appears to be a different protein (Maciejewski et al 1999).…”