Subcellular distribution of the β-N-methylamino-L-alanine-containing proteins in marine diatoms

Neurotoxin β-N-methylamino-L-alanine (BMAA) has been implicated as a major inducer of human neurodegenerative diseases. In recent years, marine diatoms were verified to produce BMAA-containing proteins. It will be an important cue to elucidate the subcellular distribution of BMAA in marine diatoms for disclosing its biosynthesis pathway. In this study, three species of Thalassiosira (T. andamanica, T. allenii and T. minima) were used to investigate the subcellular distribution of BMAA in organelles. Results showed that the crushing efficiency of diatoms was species-specific and increased with the rise of ultrasonic intensity of 22, 50 and 100 W (pulse = 0.2 s/s, 4 min), of which T. andamanica and T. allenii obtained the lowest and highest crushing efficiency, respectively. Interestingly, although T. allenii and T. minima were more efficiently crushed at 50 W and 100 W power (pulse = 0.2 s/s), their organelles were largely fragmented, which was verified by cytochrome c oxidase (CCO) enzyme analysis and transmission electron microscopy (TEM) observation. Their organelles were not fragmented only at 22 W. However, the crushing efficiency of T. andamanica was more reliable, and its organelles were essentially intact and only damaged at 100 W. Analysis of the BMAA-containing proteins showed that these proteins exclusively distribute in the Golgi and endoplasmic reticulum (ER) organelles. The nearly intact membranes of nucleus, mitochondria, Golgi and ER organelles testified that the absence of BMAA in other organelles was not caused by damage of nucleus or mitochondria. Results demonstrated that the BMAA-containing proteins were produced and accumulated in the ER and Golgi of diatoms.