RESUMEN
CD45 is a transmembrane glycoprotein possessing tyrosine phosphatase activity, which is involved in cell signaling. CD45 is expressed on the surface of most leukocytes and can be alternatively spliced by the inclusion or skipping of three variable exons (4, 5, and 6 or A, B, and C) to produce up to eight isoforms. In T cells, the splicing pattern of CD45 isoforms changes after activation; naive cells express high m.w. isoforms of CD45 which predominantly express exon A (CD45RA), whereas activated cells lose expression of exon A to form low m.w. isoforms of CD45 including CD45RO. Little is known about the specific factors controlling the switch in CD45 splicing which occurs on activation. In this study, we examined the influence of the SR family of splicing factors, which, like CD45, are expressed in tissue-specific patterns and have been shown to modulate the alternative splicing of a variety of transcripts. We show that specific SR proteins have antagonistic effects on CD45 splicing, leading either to exon inclusion or skipping. Furthermore, we were able to demonstrate specific changes in the SR protein expression pattern during T cell activation.
Asunto(s)
Empalme Alternativo/inmunología , Antígenos Comunes de Leucocito/genética , Proteínas Nucleares/fisiología , Fosfoproteínas/fisiología , Proteínas de Unión al ARN/fisiología , Animales , Arginina/fisiología , Células COS , Exones/inmunología , Humanos , Antígenos Comunes de Leucocito/metabolismo , Proteínas Nucleares/biosíntesis , Fosfoproteínas/biosíntesis , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/genética , Estructura Terciaria de Proteína/fisiología , Precursores del ARN/fisiología , Proteínas de Unión al ARN/biosíntesis , Serina/fisiología , Factores de Empalme Serina-Arginina , Linfocitos T/inmunología , Linfocitos T/metabolismo , TransfecciónRESUMEN
In acute infectious mononucleosis (AIM), very large clones of Ag-specific CD8+ effector T cells are generated. Many clones persist as memory cells, although the clone size is greatly reduced. It would be expected that the large number of cell divisions occurring during clonal expansion would lead to shortening of telomeres, predisposing to replicative senescence. Instead, we show that clonally expanded CD8+ T cells in AIM have paradoxical preservation of telomere length in association with marked up-regulation of telomerase. We postulate that this allows a proportion of responding T cells to enter the memory pool with a preserved capacity to continue dividing so that long-term immunological memory can be maintained.
Asunto(s)
Linfocitos T CD8-positivos/enzimología , Herpesvirus Humano 4/inmunología , Telomerasa/biosíntesis , Telómero , Regulación hacia Arriba/inmunología , Enfermedad Aguda , Adolescente , Adulto , Linfocitos T CD4-Positivos/citología , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/virología , División Celular/inmunología , Senescencia Celular/inmunología , Células Clonales/citología , Células Clonales/enzimología , Células Clonales/virología , Estudios de Seguimiento , Humanos , Memoria Inmunológica , Mononucleosis Infecciosa/enzimología , Mononucleosis Infecciosa/inmunología , Mononucleosis Infecciosa/patología , Receptores de Antígenos de Linfocitos T alfa-beta/fisiología , Telomerasa/fisiología , Telómero/enzimología , Telómero/inmunología , Telómero/virologíaRESUMEN
The leukocyte common antigen (CD45) is alternatively spliced, generating various isoforms expressed on hemopoietic cells. The splicing pattern of CD45 in T cells is altered in some individuals who show abnormal expression of high molecular weight isoforms containing exon A. The variant splicing pattern was shown to be associated with heterozygosity for a silent point mutation within CD45 exon A. This C to G transition is located 77 nucleotides downstream of the splice acceptor junction of exon A (198 bp total length). Here we report that this mutation is the cause of abnormal splicing. To isolate the mutant gene, somatic cell hybrids of lymphocytes with a CD45 splicing defect and a mouse lymphoid line were produced and clones expressing different isoforms of CD45 were isolated. Expression of the high molecular weight isoform containing exon A was associated with the mutation within exon A. All hybrids expressing the low molecular weight isoforms lacking exon A contained the normal allele of CD45 only. In addition, minigenes including this mutation were constructed and transfected into various cell lines (COS-7, HeLa, CHO). Semi-quantitative reverse transcription polymerase chain reaction showed an increase of more than tenfold in splicing to CD45RA (concomitant with a decrease in splicing to CD45RO) when compared with the normal minigene. Taken together, these results demonstrate a causal relationship between the mutation in CD45 exon A and the variant splicing pattern observed. The involvement of trans-acting splicing factors that interact with this region of CD45 pre-mRNA is currently under investigation.