De novo organelle biogenesis in the cyanobacterium TDX16 released from the green alga Haematococcus pluvialis

It is generally accepted that eukaryotic cell arises from prokaryotic cell, which means that organelle can be formed in prokaryotic cell. However, no such an instance has been detected till now. Here, we report organelle biogenesis in the endosymbiotic cyanobacterium TDX16 released from the green alga Haematococcus pluvialis, which occurred through six steps. (1) An inner intracytoplasmic membrane (IIM), an outer intracytoplasmic membrane (OIM) and an intervening peptidoglycan-like layer (PL) were synthesized by merging cytoplasmic membrane (CM)-derived thick margin vesicles, which partitioned the thylakoid-less cytoplasm into three compartments: an inner cytoplasm (ICP), an outer cytoplasm (OCP) and a sandwiched intracytoplasmic space (IS). (2) Osmiophilic granules that blistered from CM, OIM and IIM developed into primary thylakoids (PT) in ICP; while OCP disappeared and thus OIM and CM combined into a double-membraned cytoplasmic envelope (CE). (3) ICP decondensed; IIM and PT disassembled into tiny vesicles (TV) and double-membraned vesicles (DMV) respectively. Such that DNA fibers (DF) aggregated and migrated to PL. (4) TV fused into a double-membraned intracytoplasmic envelope (ICE), which re-compartmentalized the coalesced IS and ICP into a new intracytoplasmic space (NS) sequestering most DF and a new inner cytoplasm (NIC) with only few DF. Then ribosomes were formed in both NS and NIC, while DMV opened and extended into secondary thylakoids (ST) only in NIC. (5) NIC developed into the primitive chloroplast (PC) surrounded by ICE, in which ST disassembled, while ST-derived plastoglobuli developed into primitive eukaryotic thylakoids (PMT). After PL dismantled, the matrix of NS was concentrated and encased with the membranes synthesized from ICE-derived dotted vesicles into the primitive nucleus (PN). So, NS vanished, CE wrapped PC and PN. Outside CE, eukaryotic cell wall was formed by assembling sheath at the outer membrane of original cell wall and modifying the peptidoglycan layer. (6) Eukaryotic cytoplasm was built up from the matrix extruded from PN. Mitochondria were assembled in and segregated from PC by encapsulating a portion of stroma with the membranes synthesized from PMT-derived dense-margined vesicles. Then, most mitochondria turned into double-membraned vacuoles after matrix degradation, which mediated unconventional exocytosis and endocytosis. When this process finished, PN got matured into a nucleus enclosed by two sets of envelopes; PC matured into a chloroplast with its PMT maturing into thylakoids. Consequently, the prokaryotic TDX16 cell developed into a eukaryotic cell (TDX16-DE). Results of pigment analyses and 16S rRNA sequencing revealed that TDX16-DE chloroplast contained chlorophyll b and lutein showing 99% similarity to that of Chlorella vulgaris, TDX16 was a phycocyanin-containing cyanobacterium resembling Chroococcidiopsis thermalis (98% identity). Whereas, TDX16’s genome size (15,333,193 bp) and gene number (13,415) were 2.43 and 2.40 times those of C. thermalis (6,315,792 bp; 5593 genes) respectively, indicating that TDX16 acquired at least 7822 genes from its host H. pluvialis. Therefore, the mechanism underlying organelle biogenesis in TDX16 was the integration and expression of the obtained genes.