Pollen Tube Growth Oscillations and Intracellular Calcium Levels Are Reversibly Modulated by Actin Polymerization

Abstract

Prevention of actin polymerization with low concentrations of latrunculin B (Lat-B; 2 nM) exerts a profound inhibitory effect on pollen tube growth. Using flow-through chambers, we show that growth retardation starts after 10 min treatment with 2 nM Lat-B, and by 15 to 20 min reaches a basal rate of 0.1 to 0.2 mm/s, during which the pollen tube exhibits relatively few oscillations. If treated for 30 min, complete stoppage of growth can occur. Studies on the intracellular Ca 21 concentration indicate that the tipfocused gradient declines in parallel with the inhibition of growth. Tubes exhibiting nonoscillating growth display a similarly reduced and nonoscillating Ca 21 gradient. Studies on the pH gradient indicate that Lat-B eliminates the acidic domain at the extreme apex, and causes the alkaline band to move more closely to the tip. Removing Lat-B and returning the cells to control medium reverses these effects. Phalloidin staining of F-actin reveals that 2 nM Lat-B degrades the cortical fringe; it also disorganizes the microfilaments in the shank causing the longitudinally oriented elements to be disposed in swirls. Cytoplasmic streaming continues under these conditions, however the clear zone is obliterated with all organelles moving into and through the extreme apex of the tube. We suggest that actin polymerization promotes pollen tube growth through extension of the cortical actin fringe, which serves as a track to target cell wall vesicles to preferred exocytotic sites on the plasma membrane.

Exocytosis Precedes and Predicts the Increase in Growth in Oscillating Pollen Tubes

“…This essential ion (Brewbaker and Kwack, 1963) forms a steep tipfocused gradient at the extreme apex of the tube (Rathore et al, 1991;Miller et al, 1992;Malhó et al, 1994;Pierson et al, 1994;Roy et al, 1999) where it may stimulate exocytosis (Battey et al, 1999;Roy et al, 1999). Further studies have revealed that the Ca 2+ gradient oscillates in magnitude with the same period as growth rate; however, it follows the increase in growth rate by 10 to 408 (Messerli et al, 2000;Cá rdenas et al, 2008). Therefore, it becomes an interesting question whether or not the temporal pattern of exocytosis matches that of Ca 2+ .…”

“…Recent studies have identified several anticipatory processes. First, F-actin amount and activity increases in the apical domain in anticipation of an increase in growth rate (Fu et al, 2001;Lovy-Wheeler et al, 2007;Cá rdenas et al, 2008). Second, the alkaline band near the apex becomes more alkaline in anticipation of growth (;-808) (Lovy-Wheeler et al, 2006), presumably due to a H + -ATPase at the plasma membrane (Certal et al, 2008), whose activity oscillates.…”

“…Whereas in lily exocytosis leads growth by ;-1008, the increase in the intracellular [Ca 2+ ] follows growth by ;+10 to 408 (Messerli et al, 2000;Cá rdenas et al, 2008).…”

“…In lily (Lilium longiflorum and Lilium formosanum), pollen tube growth rates typically vary continuously from a minimum of 100 nm s 21 to >500 nm s 21 with a period of 20 to 50 s (Pierson et al, 1996;Cá rdenas et al, 2008). Additionally, many of the underlying processes also oscillate with the same period, but usually with a different phase than the growth rate (Holdaway-Clarke and Hepler, 2003;Chebli and Geitmann, 2007;Moreno et al, 2007).…”

Actin bundling in plants

“…As only tip growth, but not cytoplasmic streaming, is affected by such treatment, the subapical actin structure is likely to be closely related with growth. Recently, the effects of low dosages of Latrunculin B on pollen tube growth have been carefully revisited [Cardenas et al, 2008]. Growth inhibition was shown to be associated with a degradation of the subapical cortical fringe, as well as with a loss of cytoplasmic zonation, a diminution of the Ca 21 gradient, and a forward motion of the alkaline band.…”