Phase-specific mRNA

The phase and tissue specific puff patterns are microscopically visible patterns of active genes. However, gene activity means production of mRNA. mRNA of one particular composition should correspond to one particular pattern of active genes and mRNA of another composition to another pattern of active genes. Thus, it should be possible to demon- 

A giant chromosome from the suspensor of Phaseolus coccineus. The local loosenings of the structure correspond to the formations of puffs (from Nagl 1970). 

Now to the main experiment (Fig. 146b) using the technique outlined above, H -mRNA from two-day old cotyledons was compared with C -mRNA from 14-day old cotyledons. Great differences in the elution profiles are found, particularly in the region of rRNA and mRNA. What is of interest for us is that the mRNA from different states of seedling development are different. Thus, differential gene activities can also be demonstrated at the level of mRNA. 

---

Based on these findings and toxicology studies [233], human clinical phase I studies have been initiated [234]. In tumor biopsies from melanoma patients obtained after treatment, a reduction was found in both the specific mRNA (M2 subunit of ribonucleotide reductase, RRM2) and in the protein (RRM2) levels when compared to pre-dosed tissue. The presence of an mRNA fragment that... 

Buchner RR, Hugh TE, Ember JA, Morgan EL (1995) Expression of functional receptors for human C5a anaphylatoxin (CD88) on the human hepatocellular carcinoma cell line HepG2. Stimulation of acute-phase protein-specific mRNA and protein synthesis by human C5a anaphylatoxin. J Immunol 155 308-315. 

Bjorklund S, Skog S, Tribukait B, Thelander L (1990) S-phase-specific expression of mammalian ribonucleotide reductase R1 and R2 subunit mRNAs. Biochemistry 29 5452-5458... 

TRANSLATIONAL CONTROL Eukaryotic cells can respond to various stimuli (e.g., heat shock, viral infections, and cell cycle phase changes) by selectively altering protein synthesis. The covalent modification of several translation factors (nonribosomal proteins that assist in the translation process) has been observed to alter the overall protein synthesis rate and/or enhance the translation of specific mRNAs. For example, the phosphorylation of the protein eIF-2 affects the rate of hemoglobin synthesis in rabbit reticulocytes (immature red blood cells). 

Prokaryotes do not seem to make extensive use of control at the translational level, whereas eukaryotes use translational control much more widely. In part, translational control in eukaryotes occurs at the mRNA level. It may involve the sequestering of specific mRNAs by combining with specific mRNA-binding proteins and/or rapid degradation of mRNA so that they do not persist in inappropriate phases of the cell cycle. Other translational controls include the phosphorylation of factors involved in translation, as listed below ... 

In this way the different phases of embryogenesis are characterized by the synthesis of different, stage-specific mRNA molecules, which reflect the differential gene activity at the time. 

Fig. 146. Evidence for different mRNAs in cotyledons of the ground nut (Arachis hypogaea), which are phase-specific. (A) = control H - and Ci4-nucleic acids from cotyledons on the second day after germination. (B) = experiment H -nucleic acids from cotyledons on the second, C -nucleic acids from cotyledons on the fourteenth, day after germination. Considerable differences appear between the H - and C Aprofiles, particularly in the region of mRNA (right side of diagram) (modified from Key and Ingle 1966).

This multiprobe RPA offers the advantages of its sensitivity and capacity to simultaneously quantitate several mRNA species in a single sample of total RNA. This allows comparative analysis of different mRNA species within samples, moreover, by incorporating probes for housekeeping gene transcripts, the levels of individual mRNA species can be compared between samples. Furthermore, the assay is highly specific and quantitative owing to the RNase sensitivity of mismatched base pairs and the use of solution-phase hybridization driven toward completion by excess probe. Finally, the multiprobe RPA can be performed on total RNA preparations derived by standard methods from either frozen tissues or cultured cells. 

During translation the mRNA is read in a 5 to 3 direction and protein is made in an N-terminal to C-terminal direction. Translation relies upon aminoacyl-tRNAs that carry specific amino acids and recognize the corresponding codons in mRNA by anticodon-codon base-pairing. Translation takes place in three phases initiation, elongation and termination. 

---