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Serine transport and membrane depolarization in the liverwort Riccia fluitans.

The plasmalemma of thallus cells of the aquatic liverwort, Riccia fluitans, is reversibly depolarized by L- and D-serine. At 0.1 mM K+ in the medium, the depolarization saturates at 50 mV; half-maximal depolarization occurs at 13 microM L-serine and 30 microM D-serine, respectively. Uptake of 14C-labelled L-serine depends upon the K+ concentration and is sensitive to the membrane potential as indicated by its reduction through 1 mM sodium cyanide. We propose that serine binds to and is transported by an electrogenic carrier. However, an interaction of serine with K+ channels of the membrane seems also possible.

Biological Transport

The SPL-family transcription factor MpSPL3 orchestrates the proper regulation of vegetative and reproductive programs in Marchantia polymorpha.

SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) genes encode plant-specific transcription factors that are widely distributed across the plant kingdom. In angiosperms, the multimember SPL family regulates various biological processes, including vegetative-to-reproductive phase transition, inflorescence architecture, and lateral organ development. In contrast, the liverwort Marchantia polymorpha genome encodes only four SPL genes, with functional studies available only for microRNA-targeted members, MpSPL1 and MpSPL2. MpSPL1 was shown to control the meristem dormancy to modulate the thallus architecture, whereas MpSPL2 was found to promote the transition from vegetative-to-reproductive phase. Here, we investigate the impact of the MpSPL3 gene on M. polymorpha development. We demonstrate that MpSPL3 influences coordination of the vegetative growth and the reproductive phase transition. Knockout of MpSPL3 leads to strong growth retardation with disordered thallus morphology, reduced gemma cup number, and, most strikingly, complete loss of gametangiophore formation. Interestingly, overexpression of MpSPL3.2, the shorter isoform, has no detectable morphological effect, whereas the overexpression of MpSPL3.1, the longer isoform encoding a protein with an additional 61-aa long fragment, results in a delay in timing and reduced efficiency of gametangiophore production. Moreover, all the observed developmental abnormalities might be a consequence of the altered expression of genes essential for proper vegetative development and responsible for germ cell specification in MpSPL3 knockout and overexpression plants. Altogether, our findings demonstrate that MpSPL3 is important in regulating gametophyte development and ensuring reproductive success in M. polymorpha.

Marchantia

A study of the primary effect of the uncoupler carbonyl cyanide m-chlorophenylhydrazone on membrane potential and conductance in Riccia fluitans.

1. In the presence of 10(-5) to 10 (-8) M carbonyl cyanide m-chlorophenylhydrazone (CCCP) the membrane potential of thallus cells of the aquatic liverwort Riccia fluitans responds to changes of the external pH between 5.5 and 8.3. This occurs in the light and dark, and also if respiration is abolished by addition of 10(-4) M KCN and 10 (-5) M salicyl-hydroxamic acid. 2. The ATP-level of the thallus is reduced, independently of the external pH, by greter than or equal to 10 (-6) M CCCP to 30--40% of the control level of about 1.1 nmol ATP per mg dryweight within 5 min. 3. Upon addition of 10 (-4) M CCCP at 20 degrees C the ATP-level declines exponentially with a half time of about 20--30 s, whereas the membrane potential declines exponentially with a half time of about 2--3 s. 4. At pH 7.2 the electrical membrane conductance yields a sigmoid curve as a function of the logarithm of the CCCP concentration between 10 (-8) and 3 - 10 (-6) M. On the other hand, at 3 - 10(-7) M M CCCP the gm (electrical slope conductance, muS - cm-2 = 10 (-6)-omega-1-cm-2) versus pH-curve displays an optimum between pH 6.5 and 7.5. 5. We conclude that CCCP acts upon membrane potential and conductance in Riccia predominantly by inducing a passive proton permeability of the cell membrane, i.e. CCCP raises the permeability ratio, PH/PK, more than 100-fold above its control level of about 10.

Adenosine Triphosphate

New insights into bryophyte arabinogalactan-proteins from a hornwort and a moss model organism.

Two bryophyte models, the hornwort Anthoceros agrestis (Anthoceros) and the moss Physcomitrium patens (Physcomitrium), were analyzed for the presence of arabinogalactan-proteins (AGPs), as the emergence of these signaling glycoproteins in evolution is still under debate. AGPs of both species had a galactan core structure similar to that of other bryophyte and fern AGPs, but different from angiosperm AGPs, as 1,6-linked pyranosidic galactose was almost absent. In the Physcomitrium AGP, furanosidic arabinose (Araf) linkages were mainly terminal (10%) or 5-linked (13%), while in Anthoceros, terminal Araf dominated (26%) and was accompanied by very low amounts of 1,3-Araf and pyranosidic terminal Ara. Unusual 3-O-methylated pyranosidic rhamnose, which has never been detected in cell walls of angiosperms, occurred in both bryophyte AGPs (5% in Anthoceros, 10% in Physcomitrium AGP). This was comparable to AGPs of other spore-producing land plants. A bioinformatic search in the genomes of 14 bryophyte species revealed that most hornworts lack sequences encoding GPI-anchored classical AGPs. Generally, hornworts contained fewer sequences for AGP protein backbones compared with the liverwort Marchantia polymorpha and the moss P. patens. All of them comprise sequences for chimeric AGPs, and among those, surprisingly xylogen-like AGPs. Homologous sequences encoding glycosyltransferases and other enzymes involved in the synthesis and decoration of the AGP galactan framework were present in all bryophyte genomes. Immunocytochemistry of Anthoceros tissue detected AGPs at the plasma membrane/cell wall interface but also at the tonoplast, suggesting new functions of AGPs in bryophytes.

Mucoproteins

Effects of light on chloroplast translation in Marchantia polymorpha are similar to those in angiosperms and are not influenced by light-independent chlorophyll synthesis.

Translation of the chloroplast psbA mRNA in angiosperms is activated by photodamage of its gene product, the D1 subunit of photosystem II (PSII), providing nascent D1 for PSII repair. The involvement of chlorophyll in the regulatory mechanism has been suggested due to the regulatory roles of proteins proposed to mediate chlorophyll/D1 transactions and the fact that chlorophyll is synthesized only in the light in angiosperms. We used ribosome profiling and RNA-seq to address whether the effects of light on chloroplast translation are conserved in the liverwort Marchantia (Marchantia polymorpha), which synthesizes chlorophyll in both the dark and the light. As in angiosperms, ribosome occupancy on psbA mRNA decreased rapidly upon shifting plants to the dark and was rapidly restored upon a transfer back to the light, whereas ribosome occupancy on other chloroplast mRNAs changed very little. The results were similar in a Marchantia mutant unable to synthesize chlorophyll in the dark. Those results, in conjunction with pulse-labeling data, suggest that light elicits a plastome-wide activation of translation elongation and a specific increase in psbA translation initiation in Marchantia, as in angiosperms. These findings show that light regulates chloroplast translation similarly in vascular and non-vascular plants, and that constitutive chlorophyll synthesis does not affect light-regulated psbA translation initiation. Additionally, the translational outputs of chloroplast genes are similar in Marchantia and angiosperms but result from differing contributions of mRNA abundance and translational efficiencies. This adds to the evidence that chloroplast mRNA abundance and translational efficiencies co-evolve under selection to maintain protein outputs.

Chloroplasts