From intensive care to rehabilitation: survey on the satisfaction with care received during prolonged hospitalization for COVID-19 at a northern Italian university hospital.

OBJECTIVE: Investigating the experiences perceived by COVID-19 inpatients is a fundamental research area that is starting to be explored. For this reason, our objective was to provide the first Italian survey on COVID-19 inpatients' satisfaction, obtained through a self-completed questionnaire previously used in a reference study in a UK cohort of COVID-19 patients. SUBJECTS AND METHODS: Hospitalized COVID-19 patients (>20 days) admitted to Ferrara University Hospital who underwent rehabilitation during their hospital stay were invited to complete an anonymous questionnaire. The survey's questions explored the patients' satisfaction with the health services received, and their completion took place approximately one year after hospitalization. Information on sex, number of wards, ICU stays, and hospital discharge dates was collected. RESULTS: Sixty-two completed questionnaires were analyzed. The average overall satisfaction score obtained from the answers indicated by the participants in the tenth question was 4.7 out of 5.0. Very positive responses were observed for information about discharge plans, privacy, management of pain, sleep quality, and feeling of safety. The possibility of being consulted about medications and side effects received a very low satisfaction score. Considering overall satisfaction, no significant differences were noted for sex or ICU stay. The obtained results were almost superimposable to those reported in the cohort of COVID-19 patients of the reference study. CONCLUSIONS: This survey suggested that COVID-19 patients' healthcare satisfaction was high. Nevertheless, some areas must be improved, such as the communication and involvement of the patients in the decision-making of care and the discussion about medications or possible side effects.

Reduction of G-box binding factor DNA binding activity, but not G-box binding factor abundance, causes the downregulation of RBCS2 expression during early tomato fruit development.

The downregulation of RBCS2 promoter activity during tomato fruit development has been investigated by transient gene expression. A major drop in promoter activity occurs between 5 and 25 mm fruit diameter, corresponding to the late cell division to early cell enlargement phase. This drop is abolished by a mutation of the single G-box element necessary for high RBCS2 promoter activity in young tomato fruit. The G-box binding activity of fruit nuclear and total protein extracts drops concomitantly with the reduction of RBCS2 promoter activity while G-box binding factor expression is not affected. The data indicate that the developmental signal that downregulates the RBCS2 promoter acts on the regulation of DNA binding activity of constitutively expressed G-box binding factors.

The tomato I-box binding factor LeMYBI is a member of a novel class of myb-like proteins.

The RBCS3A gene of tomato belongs to a small gene family consisting of five members. Although the RBCS1, RBCS2 and RBCS3A promoters contain closely related cis regulatory sequences, the expression patterns of the genes are different. Whereas the RBCS1 and RBCS2 genes are expressed in both leaves and young fruit, the RBCS3A promoter is highly active in leaves, but not in young fruit. This lack of transcription could be due to a mutation in the RBCS3A promoter creating the so-called F-box, a protein binding site located between the activating cis elements, the I-box and G-box. In order to identify proteins that bind to the RBCS3A I-box/F-box region, the yeast one-hybrid system was used. One clone, LeMYBI was isolated which contains strong similarity to plant myb transcription factors. The encoded LeMYBI protein is at least 188 amino acids in length and contains two myb-like domains located at the amino terminus and close to the carboxy terminus, separated by a negatively charged domain. The protein contains a SHAQKYF amino acid signature motif in the second myb-like repeat, which is highly conserved in a number of recently identified plant myb-related genes, thus defining a new class of plant DNA-binding proteins. LeMYBI binds specifically to the I-box sequence of the RBCS1, RBCS2 and RBCS3A promoters, therefore representing the first cloned I-box binding factor. LeMYBI acts as a transcriptional activator in yeast and plants, and binds to the I-box with a DNA-binding domain located in the carboxyterminal domain.

Molecular analysis of a second functional A1 gene (dihydroflavonol 4-reductase) in Zea mays.

Some genes involved in anthocyanin biosynthesis in Zea mays are duplicated and differentially expressed. From the analysis of the A1 gene (dihydroflavonol 4-reductase), which is involved in this pathway, no molecular evidence for gene duplication was known to date. Isolation and analysis of A1 homologous genomic clones revealed the presence of a second A1 gene in maize and also two copies of the gene in Teosinte guerrero. The duplicated genes are structurally very similar and, at least in maize, the second gene is expressed.

The maize repressor-like gene intensifier1 shares homology with the r1/b1 multigene family of transcription factors and exhibits missplicing.

The recessive mutation intensifier1 of maize apparently causes an overall increase in flavonoid production in the aleurone. The mechanism by which this is achieved is not understood. We have succeeded in cloning the intensifier1 gene by transposon tagging with Suppressor-mutator and found, by sequence analyses, that it shares homology with known transcription factors in the anthocyanin pathway, in particular the r1/b1 multigene family in maize. Two cDNAs and a genomic clone were completely sequenced, and together they showed that the transcripts were misspliced. The frequency of missplicing was investigated by polymerase chain reaction analyses and sequencing of the individual introns. These studies indicate that very little functional transcript was made. Indeed, missplicing may be a mechanism for reducing the levels of a transcription factor that, when present, acts as a repressor of anthocyanin biosynthesis.

Molecular analysis of protein domain function encoded by the myb-homologous maize genes C1, Zm 1 and Zm 38.

Two maize genes, Zm 1 and Zm 38, related to the regulatory anthocyanin gene C1 were analyzed molecularly and used for fusion constructs in transient domain swapping experiments with the C1 wild-type gene. It was shown that both genes (Zm 1 and Zm 38) influence the expression of the A1 locus, a target gene for C1. Zm 1 activates the A1 promoter, however it does not turn on the whole anthocyanin pathway. The Zm 38 gene product shows functions similar to C1-I, a dominant inhibitor of the C1 wild-type gene. Concerning the trans-inhibition by C1-I two effects seem to be involved, competition for binding and formation of heterodimers. Further analysis of C1 function was carried out by a fine structure analysis of C1 mutants induced by the insertion and excision of transposable elements. These experiments indicate that for the activating domain of the protein, the formation of an alpha helix seems to be more important than a high negative charge.

Molecular analysis of C1 alleles in Zea mays defines regions involved in the expression of this regulatory gene.

The structure and function of several C1 alleles have been investigated molecularly and the importance of C1 promoter sequences for gene expression was studied using transient transformation assays. The C1 mutants analyzed were the overexpressing allele C1-S, the light-inducible allele c1-p, the null recessive allele c1-n, and the Ds element-induced allele c1-m1. Nucleotide sequence analysis of the alleles revealed a number of differences, predominantly located at the 3' end of the gene. The promoter sequences of the C1 alleles investigated so far (including wild-type and the dominant inhibitor C1-I allele) are almost identical except for two short footprint-like sequences (Box I and Box II) close to the putative CAAT box. Northern blot experiments and transient expression in particle gun experiments indicate that these sequences may be correlated with the different expression patterns of the alleles in the aleurone of maturing and germinating kernels.

The duplicated chalcone synthase genes C2 and Whp (white pollen) of Zea mays are independently regulated; evidence for translational control of Whp expression by the anthocyanin intensifying gene in.

Two chalcone synthase genes in maize have been cloned and molecularly characterized to be the C2 and the Whp (white pollen) locus. The two genes have highly homologous exon sequences but differ considerably in sequences 5' upstream and 3' downstream of the coding region, as well as in their introns. Northern and Western experiments of chalcone synthase expression in various tissues and in different genotypes indicated that C2 and Whp are differently regulated. The expression of Whp in maize aleurone is dependent on the presence of the recessive allele of the gene intensifier (in). The regulatory effect of in on Whp expression is not detectable at the transcriptional level, but seems to take place during translation.

The regulatory c1 locus of Zea mays encodes a protein with homology to myb proto-oncogene products and with structural similarities to transcriptional activators.

The structure of the wild-type c1 locus of Zea mays was determined by sequence analysis of one genomic and two cDNA clones. The coding region is composed of three exons (150 bp, 129 bp and one, at least 720 bp) and two small introns (88 bp and 145 bp). Transcription of the mRNAs corresponding to the two cDNA clones cLC6 (1.1 kb) and cLC28 (2.1 kb) starts from the same promoter. Both cDNAs are identical except that cLC28 extends further at its 3' end. A putative protein, 273 amino acids in length was deduced from the sequence of both transcripts. It contains two domains, one basic and the other acidic and might function as a transcriptional activator. The basic domain of this c1-encoded protein shows 40% sequence homology to the protein products of animal myb proto-oncogenes.

Molecular cloning of the c locus of Zea mays: a locus regulating the anthocyanin pathway.

The c locus of Zea mays, involved in the regulation of anthocyanin biosynthesis, has been cloned by transposon tagging. A clone (# 18En) containing a full size En1 element was initially isolated from the En element-induced mutable allele c-m668655. Sequences of clone # 18En flanking the En1 element were used to clone other c mutants, whose structure was predicted genetically. Clone #23En (isolated from c-m668613) contained a full size En1 element, clone #3Ds (isolated from c-m2) a Ds element and clone # 5 (isolated from c+) had no element on the cloned fragment. From these data we conclude that the clones obtained contain at least part of the c locus. Preliminary data on transcript analysis using a 1-kb DNA fragment from wild-type clone # 5 showed that at least three transcripts are encoded by that part of the locus, indicating that c is a complex locus.

The Spm (En) transposable element controls the excision of a 2-kb DNA insert at the wx allele of Zea mays.

The waxy (Wx) locus of Zea mays was cloned from strains carrying the wild-type and wx mutant alleles. The receptor component of the Suppressor-Mutator (Spm) controlling element system in the wx allele was shown to be a 2 kb long insertion within the transcribed region of the Wx gene. The insertion, termed Spm-I8, is excised during somatic reversion events induced by the autonomous controlling element Enhancer (En), which is an equivalent to Spm. Integration of Spm-I8 into the Wx gene generates a 3-bp target site duplication. Spm-I8 has a 13 bp long inverted repeat at its termini. The ends of the element can be further folded to build a large double-stranded structure consisting of five perfectly matching double-stranded regions of 9-13 bp in length, interrupted by single-stranded loops. A comparison of the wild-type and wx alleles revealed two additional insertions 6 (insert-1) and 0.25 (insert-2) kb in length. No En-induced excision of insert-1 and insert-2 could be detected so far. There is remarkable structure and sequence homology between Spm-I8 and the transposable elements Tam1 and Tam2 of Antirrhinum majus at their termini, reflecting a possible evolutionary and/or functional relationship between transposons in different plant species.

Similarity of the Cin1 repetitive family of Zea mays to eukaryotic transposable elements.

It has been suggested that the middle repetitive class of sequences that make up a large proportion of the eukaryotic genome have been amplified and dispersed by DNA transposition. Transposition is a phenomenon first postulated by Barbara McClintock on the basis of her genetic analysis of mutants in Zea mays. Since then, DNA transposition has been studied genetically in various plant systems and is well documented on the molecular level in both prokaryotes and eukaryotes. This has included the isolation of DNA inserts at various loci in several plants; however, the prevalence of transposition in plants is not established. We report here DNA nucleotide sequence data which show that some members of the Cin1 middle repetitive family of maize have features characteristic of known transposable elements. One cloned Cin1 repeat has a 6-base pair (bp) perfect inverted repeat sequence at its ends. The terminal five base pairs (5' TGTTG . . . CAACA 3') are identical to the termini of Drosophila copia transposable elements. Two other Cin1 alleles are flanked by 5-bp direct repeats. A comparison is made with the long terminal repeat (LTR) of the copia-Ty1-retrovirus families of moveable genetic elements.

Electrophoretic fractionation and translation in vitro of poly(rA)-containing RNA from maize endosperm. Evidence of two mRNAs coding for zein protein.

In the search for individual mRNAs coding for particular zein proteins, polysomal RNA was isolated from the endosperm of 22-days post-pollination maize kernels. This RNA was enriched for poly(rA)-containing RNAs and then submitted to preparative polyacrylamide gel electrophoresis under denaturing conditions. After electrophoresis, RNA fractions were eluted from the polyacrylamide gel and analyzed for zein mRNA activity by translation in vitro in the wheat germ system. The wheat germ system had previously been optimized for accurate translation of zein mRNAs. By a gel-electrophoretic analysis of immunoprecipitated products from the translation reactions in vitro, it could conclusively be shown that the endosperm of developing maize kernels contains two separable mRNAs for zein, one coding for the 22000-Mr protein and one coding for the 19000-Mr protein.

Short RNA chains synthesized at low pH are initiated at promoter sites.

Under non optimal conditions- either with limiting substrate concentrations (1) or at low pH (2)- RNA polymerase of Escherichia coli synthesizes very short RNA chains. By sequencing one RNA species synthesized at pH 5.8 upon T7 DNA we were able to demonstrate that under these conditions transcription is initiated at a normal promoter site (here A1) but however is terminated soon afterwards at specific artificial sites not used in vivo.