Lymphocyte surface modulation and cyclic nucleotides I. Topographic correlation of cyclic adenosine 3':5'-monophosphate and immunoglobulin immunofluorescence during lymphocyte capping.

Abstract

Concepts of plasma membrane structure and function have changed markedly during the past decade (1). The cell surface is no longer considered to be a rigid, static barrier between cytosol and extracellular milieu but rather a dynamic matrix in which cell surface constituents can change their relative and absolute positions (2). Lymphocyte immunoglobulin (Ig) capping is an example of this phenomenon (3, 4). Human peripheral blood B lymphocytes have surface Ig evenly dispersed on the plasma membrane which can be induced to aggregate when cross-linked by polyvalent antisera (5). The formation of local Ig patches is followed by capping which brings the Ig patches to one pole of the cell. Why or how such cross-linking of Ig results in the observed translocation is unknown. However, recent experiments have suggested the involvement of the cell's contractile (microfilaments) (3, 6) and cytoskeletal (microtubule) (7, 8) elements. The data contained in this communication indicate that cyclic adenosine 3':5'-monophosphate (cAMP) may act as one of the signals which regulate cell surface modulation. A dual immunofluorescence technique has been used to demonstrate a topographical correlation between patched and capped surface Ig labeled with rhodamine and specific cAMP immunofluorescence. Materials and MethodsSurface Immunoglobulin and cAMP Staining. Mononuclear cells were separated from peripheral blood by Ficoll-Isopaque centrifugation. The cells were washed three times in 10% (fetal calf serum (FCS)-RPMI 1640 at room temperature. Viability was assessed by trypan blue exclusion.The lymphocytes, in a concentration of 1-2 x 10 e in 0.05 ml of 10% FCS-RPMI 1640, were incubated with 0.05-0.1 ml of rhodamine-conjugated purified Ig or F(ab')2 fragments prepared from goat polyvalent anti-human immunoglobulin (9). After a i h incubation at 4°C the cells were washed three times with a total of 10 ml of cold 10% FCS-RPMI 1640. Redistribution of cell-bound

Dispersal and reformation of acetylcholine receptor clusters of cultured rat myotubes treated with inhibitors of energy metabolism.

“…After photography, a sketch of the cell was made, and its grid position was noted . After [12][13][14][15][16][17][18] examples were taken, the culture was incubated at 37°C . After 6 h, the cultures were again observed .…”

“…Patching, but not capping, occurs in the presence of energy metabolism inhibitors, suggesting that the latter, but not the former, is an active process (29,37,40). Further studies have implicated roles for Ca" (37,41), cyclic nucleotides (16), and the cytoskeleton (12,13,35,43), as well as metabolic energy, in the capping process.…”

Inhibition of Human NK Cell Cytotoxicity by Induction of Cyclic AMP Depends on Impaired Target Cell Recognition

“…However, high cAMPconcentrations seem to favour membrane mobility, as shown by Butman et al [6] with both specific antibody and concanavalin Ainduced 'capping', and as Steinberg [36] has demonstrated with cAMP-dependent phosphorylation of nascent actin chains in the presence of cAMP inducers [36]. Both cAMP and myosin can also be demonstrated underlying 'caps' induced by anti-immunoglobulin antibodies [9,32].…”

Antibody-induced changes in levels of cyclic adenosine monophosphate in leukaemic lymphocytes

“…Whether this is necessary for the activation ofadenyl cyclase, and consequent rise in cAMP, remains to be determined. Earp et al (1977) have detected by fluorescence microscopy an association between surface Ig patches and submembranous cAMP in normal lymphocytes. It is difficult to relate this localized and prolonged accumulation of cAMP to the transient surge in total cellular cAMP found by us.…”