An actin infrastructure is associated with eukaryotic chromosomes: structural and functional significance.
The presence of actin in eukaryotic nuclei, and, especially, its functional significance has not been well established. We have found that under routine immunocytochemical conditions, no actin can be detected in insect follicle cell nuclei by means of antibody (both mono- and polyclonal) or phalloidin staining. However, a pretreatment of nuclear preparations with two different endonucleases (deoxyribonuclease I or micrococcal nuclease) to remove a substantial amount of chromosomal DNA uncovers the presence of nuclear actin for both antibody and phalloidin detection. Employing the same nuclease digestion followed by antibody or phalloidin staining with squash preparations of Drosophila polytene chromosomes revealed that the nuclear actin is directly associated with the chromosomes. A strong positive signal in the polytene chromosomes obtained with phalloidin labeling not only confirmed the presence of actin in the chromosomes, but indicates that a considerable amount of nuclear actin is present in filamentous form (F-actin) rather than monomeric (G-actin). The detection of actin associated with Xenopus embryo chromosomes suggests the significance of chromosomal actin for diploid vertebrate cells. Using the specific actin disrupting agent cytochalasin D, we have demonstrated the structural significance of nuclear actin in maintaining the linear integrity of polytene chromosomes. Further, we present evidence that RNA polymerase II closely interacts with the chromosomal actin scaffold, and that its association with chromosomes does not require the presence of DNA.
Sauman
I
I
Wesleyan University, Biology Department, Middletown, CT 06459-0170.
Berry
S J
SJ
D000199:Actins / Q000235:genetics*; D000546:Amanitins; D000818:Animals; D002467:Cell Nucleus / Q000737:chemistry*; D002875:Chromosomes; D015638:Cytochalasin D / Q000494:pharmacology; D004331:Drosophila melanogaster / Q000235:genetics*; D004720:Endonucleases; D007124:Immunoenzyme Techniques; D009036:Moths / Q000235:genetics*; D012319:RNA Polymerase II; D013194:Staining and Labeling; D013329:Structure-Activity Relationship; D014981:Xenopus / Q000235:genetics*
D000199:Actins; D000546:Amanitins; D015638:Cytochalasin D; C018358:phallotoxin; D012319:RNA Polymerase II; D004720:Endonucleases