Temperature-sensitive formation of chloroplast protrusions and stromules in mesophyll cells of Arabidopsis thaliana

Abstract

In leaf mesophyll cells of transgenic Arabidopsis thaliana plants expressing GFP in the chloroplast, stromules (stroma-filled tubules) with a length of up to 20 mum and a diameter of about 400-600 nm are observed in cells with spaces between the chloroplasts. They appear extremely dynamic, occasionally branched or polymorphic. In order to investigate the effect of temperature on chloroplasts, we have constructed a special temperature-controlled chamber for usage with a light microscope (LM-TCC). This LM-TCC enables presetting of the temperature for investigation directly at the microscope stage with an accuracy of +/-0.1 degrees C in a temperature range of 0 degrees C to +60 degrees C. With the LM-TCC a temperature-dependent appearance of chloroplast protrusions has been found. These structures have a considerably smaller length-to-diameter ratio than typical stromules and reach a length of 3-5 mum. At 5-15 degrees C (low temperatures), almost no chloroplast protrusions are observed, but they appear with increasing temperatures. At 35-45 degrees C (high temperatures), numerous chloroplast protrusions with a beaklike appearance extend from a single chloroplast. Interaction of stromules with other organelles has also been investigated by transmission electron microscopy. At 20 degrees C, transverse sections of stromules are frequently observed with a diameter of about 450 nm. A close membrane-to-membrane contact of stromules with the nucleus and mitochondria has been visualised. Golgi stacks and microbodies are found in the spatial vicinity of stromules. At 5 degrees C, virtually no chloroplast protrusions or stromules are observed. At 35 degrees C, chloroplast protrusions are present as broader thylakoid-free stroma-filled areas, resulting in an irregular chloroplast appearance.